Method of thermoregulation within an incubator for babies before implantation

ABSTRACT

The invention relates to an incubator for babies. It is of special value for the treatment of premature infants as an intensive care unit at any time during life from creation to implantation. A cradle is sided with ports to enable fluidic ventilation. Advantageously the incubator includes an optical path for imaging the patient via a clear bottom and open top. The incubator is provided with easy access and various accessories required for an intensive care unit. The invention further relates to a method of thermoregulation within the incubator and to a temperature measurement test pattern to calibrate a radiometer for non-contact patient thermometry. Within the incubator, the temperature of a human embryo or hatchling patient is monitored distinctly from incubator temperature, and a rate of flow of a liquid incubation medium over the patient&#39;s egg (embryo stage) or body (hatchling stage) is controlled by a microfluidic means which is responsive to feedback from the patient&#39;s temperature in view of an effect of flow-related heat dissipation on the difference between patient temperature and the temperature of the liquid incubation medium bathing the patient in the incubator.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.11/549,972, filed Oct. 16, 2006, which is a continuation-in-part of U.S.application Ser. No. 10/908,861, filed May 30, 2005, now U.S. Pat. No.7,121,998, which claims the benefit of U.S. Provisional Application No.60/577,958, filed Jun. 8, 2004, which are incorporated here by way ofreference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

My invention relates to infant incubators.

The purpose of this teaching is to add new matter to the parentapplication (Ser. No. 10/908,861).

My invention relates to a vented microcradle for use in an incubatorsystem specifically designed to maintain a premature infant in acontrolled care environment from creation to implantation. The incubatorsystem is thus an engineered environment specifically designed for thecare of human embryos and hatchlings.

A human infant is properly termed an embryo only from creation untilhatching, and then from hatching until implantation he or she is termeda hatchling. Hatching is a milestone of human development in which theembryo makes a hole in the shell of the human egg and then escapes in anextrusive behavior. Hatching is a prerequisite to implantation.Incorrect definitions of the term embryo have persisted due to ignoranceof the human hatching event.

Nidation is another word for implantation. A human infant is termed aprenid (PRE-nid) from creation to implantation. The word prenid isderived by shortening of pre-nidation, which means pre-implantation, andprenidial (pre-NID-e-al) is the adjectival form. The term prenid isconvenient because it encompasses both human embryos and hatchlings.According to this etymology, the uterine cavity is the nest (Latinnidus, from which the term nidation is derived) and the settlement theinfant makes in the nest at implantation time is called the nidia(compare Latin colonus, colonia). Hence, prenidal (pre-NIGH-dal) wouldmean before the infant has entered the uterine cavity, whereas prenidialmeans before the infant has actually implanted.

Prenidial gestation refers to a maternal, bodily provision for prenidialdevelopment as well as to prenidial developmental needs in general,whereas prenidial incubation refers to an engineered provision fordevelopment outside the maternal body in a manner analogous to naturalgestation. Whether due to preterm delivery or external creation, outsidethe maternal body prenids are premature infants because it is prematurefor them to be outside the maternal body on their own.

Sophisticated incubator systems for premature infants prior toimplantation are termed prenidial incubators. Prenidial incubators areanalogous to neonatal incubators, except they relate to patient care forprenids instead of for neonates.

My invention relates to a vented microcradle for a prenidial incubator,more specifically a side-vented microcradle for a prenidial incubator.

2. Prior Art

Historically and to the present, medicine has had trouble understandingthe principles of thermoregulation needed to care for infants inincubators. Time and again practitioners have made the basic mistake ofconfusing a patient's temperature with the ambient temperature of theenvironment inside the incubator. To finally be clear on this, it isincorrect to measure the quantity corresponding to the temperatureinside an incubator and to interpret it as data corresponding to thevariable of the patient's own temperature! Instead, the patient'stemperature must be monitored distinctly in contrast to the temperatureof the patient's environment.

Another key ingredient for incubator competence is ventilation. Astagnant environment is not healthy and so a circulation of theenvironment is necessary. Though similar in principle, neonates andprenids differ in their ventilation requirements because neonates livein a gas/vapor phase environment whereas prenids live in a liquid phaseenvironment. Prenids transfer all metabolic resources and wastes via anincubation medium in which they are submerged. Prenids require gentlefluidic ventilation, which means the incubation medium must becirculated around the infant to remove wastes and to refresh metabolicresources, but gently enough so that beneficial substances produced bythe infant are not stripped away. Fluidic ventilation is provided in thefallopian tube by cilia and small muscular contractions that urge fluidto-and-fro past the infant.

My teaching in U.S. Pat. No. 6,694,175 introduces the competent mannerof thermoregulation for infants in a prenidial incubator. My teaching inthe parent application (Ser. No. 10/908,861) introduces the competentmanner of ventilation for infants in a prenidial incubator. Theseteachings are incorporated here by way of reference. These teachingscombine to establish prenidial incubation as a competent discipline forthe care of infants prior to implantation. A thorough understanding ofthese teachings is prerequisite to the present disclosure. Thoroughunderstanding of the discussions presented in papers found in the filewrappers of the corresponding applications, with particular emphasis ondiscussions of the prior art, is strongly recommended.

Following upon the parent disclosure (Ser. No. 10/908,861), thisdisclosure adds variety and complexity to the medical science ofprenidial ventilation. In addition, because the medical and scientificcommunities continue to struggle helplessly with even the most basicconcepts, the principles of prenidial thermoregulation are reviewed.

Genifection (JEN-ih-feck-shun) is the crime of using assistedreproductive technologies to create human embryos in an environmentwhere they are unlikely to be cared for with full responsibility asindividual patients, beloved family members, and equal members ofsociety. The word is coined using Latin roots, by combining geni- (one'skind) with -fection (the making of, by artful means). Genifection is aterrible crime against humanity.

At present practitioners of in vitro fertilization rely heavily ongenifection to compensate for their extremely poor success in incubatinghuman embryos and hatchlings. Unable to reduce the tragic mortalityrates that result from their poorly skilled incubation and transfermethods, practitioners of in vitro fertilization typically create aplurality of embryos so that the product of their numbers despite lowsurvival rates will sometimes result in at least one survivor for thesake of entrepreneurial success. In other words, such practitioners usegenifection to compensate commercially for their extremely poor medicaland scientific practices.

In other cases, though comparatively less common, human embryos havebeen created in the lab with only their harm in mind, e.g., forexperiments in stem cell research.

Please join me in putting an end to these dehumanizing crimes ofgenifection.

3. Statement of the Necessity

For a neonatal incubator, the cradle portion of the incubator must allowfor easy access to a patient while at the same time affording properthermoregulation and ventilation at all times. A well-designed prenidialincubator should offer the same advantages.

It may also be desirable to visualize an infant in a prenidial incubatorfrom below as well as from above. A variety of vented microcradle isneeded to satisfy this objective.

What is needed is a side-vented microcradle for a prenidial incubator,more specifically a side-vented microcradle with a clear bottom and anopen top.

4. Note

A number of cited references were found after filing the originalspecification in Ser. No. 11/549,972. These are discussed in severalpapers available in the electronic file wrapper of that application.These are available online at the U.S. Patent and Trademark Officewebsite using the public Patent Application Information Retrieval system(Public PAIR). Engineers should consider these papers along with theassociated references. Note that the non-patent literature cited isavailable in the file wrapper under “NPL Documents”.

Of particular note, Matz (U.S. Pat. No. 2,062,468), Le Pesant et al(U.S. Pat. No. 4,569,575), and Tuckerman et al (U.S. Pat. No. 4,450,472)provide teachings in their respective arts that pre-date prior artreferences cited in the original specification of Ser. No. 11/549,972.

Referring to FIG. 105, Matz teaches a variable focus liquid lens; incontrast, teachings such as those of Berge et al (U.S. Pat. No.6,369,954) and Feenstra et al (U.S. Pat. No. 7,126,903) providerelatively modern examples of renewed interest in this art.

Referring to FIG. 106, Le Pesant et al teach a digital microfluidicsystem, and refer to an even earlier example of this art; in contrast,teachings such as that of Pamula et al (U.S. Pat. No. 6,911,132) providerelatively modern examples of renewed interest in this art.

Tuckerman et al teach a use of microfluidics to cool integratedcircuits.

Also of particular note, Petronis et al describe a microfluidic cellculture chip employing substantially planar layers laminated together.However, they do not employ an open-top microcradle design. (Petronis etal, “Transparent Polymeric Cell Culture Chip with Integrated TemperatureControl and Uniform Media Perfusion,” BioTechniques, vol. 40, no. 3, pp.368-76, March 2006.) Noted is the abstract and FIG. 1, p. 369.

Kricka et al (U.S. Pat. No. 5,296,375) teach a device made ofmicromachined glass layers comprising separate wells for holding spermand egg separated by a swim-up channel for sperm. They employ pumps orcapillarity to fill channels and wells and means such as a pump orsyringe to expel a fertilized egg. (column 3, lines 51-64) They do notteach or fairly suggest fluidic ventilation of an embryo or hatchling.

BRIEF SUMMARY OF THE INVENTION

My invention satisfies the above-stated needs.

The invention comprises a microfabricated cradle (“microcradle”) formingan enclosure for the care of a premature infant during preimplantationdevelopment with a clear bottom and an open top and having one or moreside vents for fluid entry/exit to provide a circulation of fluid withinthe cradle via a system of microfluidic channels and related devices.The invention is therefore a side-vented microcradle with a clear bottomand an open top.

The microcradle along with its related systems and devices arecollectively referred to as an incubator, more specifically a prenidialincubator; similarly, a medical care environment provided by theincubator for infant care is called the incubator environment. Thepurpose of the incubator is to sustain the life of a premature infantduring life before implantation.

The invention further comprises a fluidic circuit assembly (FCA), whichis necessary for the practice of elaborate embodiments, a temperaturebath to maintain an ambient temperature for the incubator,machine-readable indicia or markings, and a specialized catheter totransfer a patient to/from the incubator.

The invention incorporates and is compatible with various modificationsof my teaching in U.S. Pat. No. 6,694,175 to achieve properthermoregulation of the patient. The invention may also incorporateadditional technologies to achieve increasing sophistication.

The invention forms a central structure in an incubator for the care ofpremature infants during life at any time prior to implantation.

The invention finds use whenever an infant requires incubation outsidethe maternal body during life before implantation.

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWING

FIG. 1 is a side cross-sectional view of a side-vented microcradle.

FIG. 2A is a side cross-sectional view of a glass petri dish.

FIGS. 2B-C are bottom orthogonal views of a glass sheet havingmicrofluidic channels and vias etched in it for a side-ventedmicrocradle.

FIG. 2D is a side cross-sectional view of a side-vented microcradle.

FIG. 3 is a side cross-sectional view of a petri dish formed into adouble-walled vessel with an inlet and outlet for a running fluid.

FIG. 4 is a side cross-sectional view of a side-vented microcradle, andis most descriptive of the invention.

FIG. 5 is an exploded perspective view of a fluidic circuit board havinga side-vented microcradle.

FIGS. 6A-D are side cross-sectional views of side-vented microcradles.

FIGS. 7A-B are bottom orthogonal views of glass sheets havingmicrofluidic channels and vias etched in them for a side-ventedmicrocradle.

FIG. 8 is a side cross-sectional view of a chamber etched in a layer tosupport a device inside a fluidic circuit board according to theinvention.

FIG. 9A-C, E are side cross-sectional views of sections of a fluidiccircuit board according to the invention.

FIG. 9D is a top orthogonal view of a fluidic circuit board having aside-vented microcradle.

FIG. 10 is a side cross-sectional view of a fluidic circuit boardaccording to the invention.

FIG. 11 is a perspective view showing steps in the manufacture of atransfer catheter formed by a fluidic circuit board according to theinvention.

FIG. 12 is a side cross-sectional view of a prior art transfer catheter.

FIG. 13 is a side cross-sectional view of a transfer catheter having aside-vented microcradle.

FIG. 14 is a perspective view of a transfer catheter according to theinvention.

FIG. 15 is a side cross-sectional view of a side-vented microcradle.

FIG. 16 is a mathematical formula relying on notation used in FIG. 15.

FIGS. 17-18 are exemplary mask patterns used in etching microfluidicchannels and vias for a side-vented microcradle.

FIG. 19 is a bottom orthogonal view of a glass sheet having microfluidicchannels and vias etched in it for a side-vented microcradle.

FIG. 20A is a side cross-sectional view of a temperature-detectinginfrared camera attached to a microscope lens to detect the bodytemperature of a patient inside a prenidial incubator according to U.S.Pat. No. 6,694,175.

FIG. 20B is a side cross-sectional view of a prior art set up forordinary microscopy of a prenid in an incubator.

FIG. 20C is a side cross-sectional view of a prior art setup for highresolution microscopy of a specimen in a petri dish using an invertedmicroscope having a high numerical aperture objective lens.

FIG. 21A is a side cross-sectional view of a temperature-controlled bedaccording to the invention.

FIG. 21B is a top cross-sectional view of the bed shown in FIG. 21A.

FIG. 22 is a side cross-sectional view of a temperature-controlled bedaccording to the invention incorporating a fluidic circuit boardassembly layer.

FIG. 23 is a side cross-sectional view of a side-vented microcradle.

FIG. 24A is a side cross-sectional view of a side-vented microcradletaken along a line 99 with respect to FIGS. 24B-C.

FIGS. 24B-C are bottom orthogonal views of glass sheets havingmicrofluidic channels and vias etched in them for the side-ventedmicrocradle shown in FIG. 24A.

FIGS. 25-26 are side cross-sectional views of side-vented microcradles.

FIG. 27 is a bottom orthogonal view of a glass sheet having microfluidicchannels and vias etched in it for a side-vented microcradle.

FIG. 28 is a side cross-sectional view of a section of a side-ventedmicrocradle.

FIGS. 29A-B are side cross-sectional views of a prior art variable focusliquid lens.

FIGS. 30A-B are side cross-sectional views of a variable focus liquidlens according to the invention.

FIGS. 31A-B are respective top and bottom orthogonal views of a glasssheet having microfluidic channels and a via etched in it for the liquidlens shown in FIGS. 30A-B.

FIG. 31C is a top orthogonal view of a glass sheet having a conductivetrace deposited on it to form an electrode for the liquid lens shown inFIGS. 30A-B.

FIG. 32 is a side cross-sectional view of a variable focus liquid lensaccording to the invention.

FIGS. 33A-B are side cross-sectional views of a variable focus liquidlens according to the invention.

FIG. 34A is a side cross-sectional view of a variable focus liquid lensaccording to the invention.

FIG. 34B is a side cross-sectional view of a number of lens shapesachievable by the liquid lens shown in FIG. 34A.

FIG. 35 shows designated top and bottom orthogonal views of glass sheetshaving microfluidic channels and vias etched in them for the liquid lensshown in FIGS. 34A-B.

FIG. 36 shows a side cross-sectional view of a variable focus liquidlens according to the invention.

FIGS. 37A-B are side cross-sectional views of lens shapes achievable bythe liquid lens shown in FIG. 36 according to alterations of electricalpolarity.

FIGS. 38A-B are side cross-sectional views of variable focus liquid lensobjectives according to the invention.

FIG. 39 is a side cross-sectional view of a prior art microscopy setupillustrating, in view of FIG. 20C, a relationship between workingdistance and numerical aperture.

FIG. 40 is a side cross-sectional view of a prior art microscopy setupillustrating a problem of lost light when employing a flat cover glass.

FIG. 41 is a side cross-sectional view of a microscopy setup employing amicrolens cover glass according to the invention.

FIG. 42 is a side cross-sectional view of a section of a side-ventedmicrocradle with optical equipment separate from its fluidic circuitassembly placed underneath.

FIG. 43 is a top orthogonal view of an array of optical devices in alayer of a fluidic circuit assembly according to the invention.

FIG. 44 is a top orthogonal view of an array of variable focus liquidlens objectives according to the invention.

FIG. 45 is a side cross-sectional view of a variable focus liquid lensobjective according to the invention.

FIG. 46 is a side cross-sectional view of a liquid lens with variableoptical properties according to the invention.

FIGS. 47A-B are side cross-sectional views of a liquid lens withvariable optical properties according to the invention.

FIGS. 48A-E are side cross-sectional views of a human infant and his orher egg capsule during different stages of prenidial life.

FIG. 49 is a graph of the electromagnetic spectrum.

FIGS. 50A-B list data on the electromagnetic spectrum.

FIG. 51 is a graph of a Planck's law distribution.

FIGS. 52A-B are graphs of electromagnetic absorption by water.

FIG. 53 is a graph of an external transmission report for sapphire.

FIG. 54A is a side cross-sectional view of a prenid in an incubator whois resting on a flooring layer.

FIG. 54B is a side cross-sectional view of a prenidial patient in anincubator who is resting on a concave microlens flooring layer accordingto the invention.

FIG. 55 is a top orthogonal view of an emissivity pattern.

FIG. 56 is a top orthogonal view of an emissivity pattern according tothe invention.

FIGS. 57A-B are graphs of transmission reports for coating substancesassociated with respective areas of the emissivity pattern shown in FIG.56.

FIGS. 58A-B are graphs comparing radiation amount as a function oftemperature for the coating substances associated with the emissivitypattern shown in FIG. 56.

FIG. 59 is a flowchart representing search criteria for finding opticalliquids to form a liquid lens.

FIG. 60 is a graph of infrared absorption for thin water films.

FIGS. 61A-B are simplified top cross-sectional views of microfluidicsystems for exchanging parcels of fluid between slow and fast movingchannels so as to speed fluid transit and delivery between a patient anda fluidic circuit assembly device.

FIGS. 62A-B are side cross-sectional views of ions and polar moleculesgathered near a sidewall electrode to illustrate a theory of capacitivespreading.

FIG. 63 is a side cross-sectional view of a prior art digitalmicrofluidic system.

FIG. 64 is a side cross-sectional view of a prior art digitalmicrofluidic system.

FIG. 65A is a top orthogonal view of an arrangement of electrodes forself-scooting circuitry according to the invention.

FIG. 65B is a side cross-sectional view of a digital microfluidic systememploying the electrode arrangement shown in FIG. 65A.

FIG. 66 is a schematic view of an electronic circuit for theself-scooting circuitry of the digital microfluidic system shown inFIGS. 65A-B.

FIGS. 67A-B, D are schematic views of electronic circuits forself-scooting circuitry according to the invention.

FIG. 67C is a formula for an electronic circuit for self-scootingcircuitry according to the invention.

FIG. 67E is a top orthogonal view of control electrodes being enabledand disabled according to the invention so as to route droplet movementat a fork.

FIG. 67F is a schematic view of a droplet switch being employedaccording to the invention to enable other circuitry.

FIG. 68 is a top orthogonal view of a liquid flowing continuously from areservoir according to the action of electrowetting.

FIGS. 69A-C are timing diagrams for self-scooting circuitry according tothe invention, showing different alternatives for turning controlelectrodes on and off over time.

FIGS. 70A-B are side cross-sectional views of a droplet sandwichedbetween electrodes to illustrate a theory of dielectric electrowetting.

FIG. 71 is a side cross-sectional view of a digital microfluidic systememploying charged electrowetting according to the invention.

FIG. 72 is a side cross-sectional view of a digital microfluidic systememploying dielectric electrowetting according to the invention.

FIG. 73 is a side cross-sectional view of a digital microfluidic systememploying charged electrowetting according to the invention.

FIG. 74 is a side cross-sectional view of an open-top digitalmicrofluidic system employing charged electrowetting according to theinvention.

FIG. 75 is a side cross-sectional view of a droplet supported by a trackof control electrodes, illustrating the effect of coulombic forces in anopen top arrangement.

FIGS. 76A-77B are top (76B, 77B) and bottom (76A, 77A) orthogonal viewsof electrode arrangements for transistorless self-scooting circuitryaccording to the invention.

FIG. 78 is a side cross-sectional view of a digital microfluidic systememploying the transistorless self-scooting circuitry shown in FIGS.76A-77B.

FIG. 79A-B are schematic views of electronic circuits according to theinvention for the transistorless self-scooting circuitry of the digitalmicrofluidic system shown in FIGS. 76A-78.

FIG. 80 is a top orthogonal view of an arrangement of electrodes forself-scooting circuitry employing a pacer track and a free trackaccording to the invention.

FIG. 81 is a schematic view of an electronic circuit for theself-scooting circuitry of the digital microfluidic system shown in FIG.80.

FIG. 82 is a top orthogonal view of an electrode arrangement forself-scooting circuitry according to the invention.

FIGS. 83A-B are side cross-sectional views of electrode arrangements forself-scooting circuitry according to the invention.

FIG. 84 is a top orthogonal view of a prior art interdigitated electrodearrangement.

FIG. 85 is a side cross-sectional view of an electrode arrangement forself-scooting circuitry according to the invention.

FIG. 86 is a side cross-sectional view of a combination lateral andvertical airflow system according to the invention.

FIG. 87 is a side cross-sectional view of a side-vented microcradle.

FIG. 88 is a floor plan for a “fertility intensive care unit” clean roomaccording to the invention.

FIGS. 89A-D are top orthogonal views of patterns of openings for fluidin a vented flooring for a microcradle according the invention and itsparent teaching (Ser. No. 10/908,861).

FIG. 90 is a side cross-sectional view of a side-vented microcradleillustrating a method according to the invention, whereby a patient islifted by fluid flow.

FIG. 91 is a top orthogonal view of a prior art compartmentalizedstructure for embryos covered by a microdrop that is stabilized bypickets.

FIGS. 92A-C are top orthogonal views of stabilizing structures for amicrodrop according to the invention; the microdrop covers a ventedmicrocradle.

FIG. 93 is a side cross-sectional view of a side-vented microcradleemploying an optical device according to the invention for sensingmicrodrop volume.

FIG. 94A is a top orthogonal view of a side-vented microcradle.

FIG. 94B is a side cross-sectional view of the side-vented microcradleshown in FIG. 94A, taken along a line 224.

FIGS. 94C-E are side cross-sectional views of a sidewall section of theside-vented microcradle shown in FIG. 94A, taken along a line 225; eachshows a different arrangement of ventilation ports in the sidewall.

FIGS. 95A-B are side cross-sectional views of micro-canals according tothe invention, which are embodiments of a side-vented microcradle.

FIG. 95C is a top orthogonal view of a micro-canal according to theinvention.

FIG. 96A is a top orthogonal view of a side-vented microcradle.

FIG. 96B is a side cross-sectional view of a side-vented microcradle.

FIG. 97 is a side cross-sectional view of a digital microfluidic systememploying self-scooting circuitry according to the invention.

FIGS. 98A-B are side cross-sectional views of a digital microfluidicsystem employing self-scooting circuitry according to the invention.

FIG. 99 is a side cross-sectional view of a digital microfluidic systememploying self-scooting circuitry according to the invention.

FIG. 100 is a top orthogonal view of an electrode arrangement for thedigital microfluidic system shown in FIG. 99.

FIG. 101 is a side cross-sectional view of a prenidial infant depictedin reference to points A, B, and C, respectively located inside thepatient's body, on the outer surface of the body, and at a distance of afew microns away from the outer surface of the body in the surroundingfluid of the incubation medium.

FIGS. 102A-103B are graphs of temperature as a function of distance frompoint B on the outer surface of the body to point C located a distanceaway in the surrounding fluid incubation medium, as designated in FIG.101.

FIG. 104 is a flowchart representing responses to chilled and overheatedpatients according to the invention.

FIG. 105 is a side cross-sectional view of a prior art variable focusliquid lens.

FIG. 106 is a side cross-sectional view of a prior art digitalmicrofluidic system.

DETAILED DESCRIPTION OF THE INVENTION

Analogous to a cradle for neonatal care, a vented microcradle forms acentral structure in an incubator environment for the care of infantsbefore implantation. As with the cradle of a neonatal incubator, keyingredients of the vented microcradle are easy access to a patientcombined with proper thermoregulation and ventilation at all times.Disclosed is a side-vented embodiment of a microcradle with a clearbottom and an open top.

The invention relies on integrated microfabrication technology (IMT) forits manufacture. IMT is a broad, interdisciplinary field employingdiverse arts to make small size systems and devices. IMT technologyincludes submillimeter, micrometer (micron), submicron, and nanometertechnologies, and often incorporates these with larger scaletechnologies. In the world of micromanufacturing, IMT designs offerimportant benefits such as intimate integration with electroniccircuitry and the ability to manufacture arrays of designs on a singlesubstrate. Although many other IMT arts are also implicated by theinvention, microfluidic technology and glass microfabrication technologyare especially relevant. Microfluidic technology relates to fluid flowon a microscopic scale and includes a number of rapidly evolvingspecialty areas such as micropump and microvalve technology. Using IMTtechnologies for prenidial incubation enables infants to be preciselyhandled and cared for in a way that is tailored to their microscopicsize and gentle fluidic requirements during life before implantation.

1. Preferred Embodiment

My invention is a side-vented microcradle with a clear bottom and anopen top for the purpose of prenidial infant care in an incubatorsetting. According to the preferred embodiment shown in FIG. 1, theside-vented microcradle has the following minimum features: a cradle 1for cradling a prenidial patient P, the cradle 1 having an open top 2, aclear bottom 3, and one or more side vents 4 to allow for an entry/exitof a fluid medium M into/out of the cradle 1 via microfluidic channels5. The fluid is a liquid incubation medium M in which the patient P issubmerged. In operation, fluid exiting the side vents 4 is translatedinto a vertical direction so as to ventilate the patient P inside thecradle 1. Similarly, fluid inside the cradle 1 may be withdrawn throughthe side vents 4 so as to ventilate the prenid P. Accordingly, fluid Mmay be made to flow over the prenid's body P in an upward, downward, orto-and-fro direction. Using microfluidics, including means to urge fluidto flow (such as means of a micropump), fluid M is urged via themicrofluidic channels 5 through the side vents 4 so as to cause fluid Mto circulate gently over the prenid's body P for the purpose of fluidicventilation.

The open-top design offers a number of distinct advantages over theprior art. Unlike the art of Hunter (U.S. Pat. No. 4,574,000), Beebe etal (U.S. Pat. No. 6,193,647), and Thompson et al (U.S. Pat. No.6,673,008), as well as what has been disclosed by Campbell et al (USpublished application 2002/0068358), using my invention the patient iseasy to access in a direct fashion, fluidic ventilation does not need tobe interrupted while the patient is being accessed, and the patient isexposed to an ambient pressure above the microcradle rather than to apressure used to urge fluid to flow.

As will appreciated by those skilled in microfluidics and other arts inthe field of integrated microfabrication technology (IMT), numerousvariations of the FIG. 1 embodiment are easy and cost-effective tomanufacture.

2. Example of Manufacture

A side-vented microcradle is preferably made of glass. Here a minimalembodiment is described to give a basic manufacturing example. Referringto FIGS. 1 & 2A-D, the clear bottom 3 of the microcradle is formed bythe bottom of a clear petri dish 6 made of glass. A desired pattern ofmicrofluidic channels 5 is etched on one side of a glass sheet 7 that inthis example is 100 microns thick. A hole 8 that in this example is 500microns in diameter is etched through the glass sheet 7. This hole 8forms the walls 9 of the cradle 1. A second hole 10 is made through theglass sheet 7 to form a via (through-hole) in fluidic communication withthe microfluidic channels 5. The glass sheet 7 is directly bonded(preferably without adhesive) to the bottom of the petri dish 6 with thepattern of microfluidic channels 5 facing the petri dish 6. Amicrofluidic connector 11 is bonded on top of the glass sheet 7 over thesecond hole 10 by melting a fine strand of glass where the edges of theconnector 11 meet the glass sheet 7. A fluid supply line 12 is connectedto the microfluidic connector 11, said line 12 being further connectedin series to a flow meter 13, micropump 14, and fluid reservoir 15. Theflow meter 13 may be omitted if the micropump 14 is self-metered oranother means of monitoring flow is provided.

To achieve operation the petri dish 6 is partially filled with a fluidincubation medium M and the microfluidic system comprising the reservoir15, micropump 14, flow meter 13, and microfluidic channels 5 is primedwith the same fluid; the micropump 14, which is preferably reversible,is operated as desired with the aid of flow rate indications. Movementof fluid in the to-and-fro direction, with net movement in onedirection, is preferred. For example, the net movement may occur in thedownward direction, meaning with fluid flowing out of the microcradlethrough the side vents 4. In this simplistic example, the fluid fillingthe petri dish 6 provides a reservoir of fresh medium M for the patient;a to-and-fro movement helps to maintain endogenously produced substancesaround the patient's body; at the same time, a net downward movementremoves wastes and draws upon fresh fluid medium M from above thepatient P. The to-and-fro movement may be likened to the beating ofcilia in the fallopian tube. It is contemplated that in order tomaintain a gentle state of fluid flow the rate of flow past the patientP should generally not exceed an upperbound of 1 to 10 microns persecond. The side vents 4, which are ventilation ports located on thesidewalls 9 of the microcradle, are formed where the microfluidicchannels 5 meet the hole 8 forming the cradle walls 9; at the pointwhere the microfluidic channels 5 meet the hole 8 forming the cradlewalls 9, the channels are 20 microns high and 150 microns wide in thisexample. Correspondence between a rate of fluid flow through the sidevents 4 and a desired flow rate past the patient P can be determined bycalculation. More detail on this topic is given under “DesignConsiderations” below. An embryo P with an eggshell 140 microns in outerdiameter will stick out a ways above the 100 micron height of the cradlewalls 9 in this example; however, in general the cradle walls can bemade to any desired height.

Referring to FIG. 3, the petri dish 6 is preferably formed into adouble-walled vessel 16 with an inlet 17 and outlet 18 for a runningfluid 19. The fluid serves as a precisely controlled temperature bath tomaintain ambient temperature for the incubator environment. Referring toFIG. 4, a microscope 20 may be positioned below to visualize the patientP and a thermal imaging camera 21 according to U.S. Pat. No. 6,694,175may be placed above to monitor the temperature of the patient P. A lamp22 used with the microscope for lighting should not produce infraredradiation at a level that will overheat the patient P; if necessary, asurface half-mirrored with gold or other filter for infrared radiationmay be used. According to U.S. Pat. No. 6,694,175, infrared heat lamps23 may be focused on the patient P from any location (here shown above)to provide warmth. More detail on the topic of positioning a thermalimaging camera and infrared heat lamps is given under “DesignConsiderations” below, particularly in reference to an absorption ofinfrared radiation by an aqueous medium. If necessary, the lamps 23 maybe shuttered each moment a thermal image is taken so that theirsignature does not interfere. The incubator is kept in a clean roomenvironment and a laminar flow hood provides additional protection fromcontaminants. Humidity, air temperature, air composition, and airpressure are maintained in the flow region of the hood.

As a notable feature, when no sources of impurity are introduced by themanufacturing process, the patient will only be exposed to the glass ofthe incubator, which is inert. Thus, this particular manufacturingexample results in an ultrapure incubator environment for the patient.Impurities associated with the manufacture of devices located down theline 12, e.g., the micropump 14, will not reach the patient if they arenot made to flow up the line 12 and into the microcradle where thepatient P rests. A net downward flow (draining fluid out of themicrocradle) offers this advantage. The ultrapure environment eliminatesa need for clinical studies on the effect of patient exposure tomanufacturing impurities, e.g., from adhesives. This explains why thedirect bonding of glass to glass with no adhesive is preferred.

Microchannels can be etched in glass using a variety of microfabricationtechnologies, including laser etching, powder blasting, DRIE (deepreactive ion etching), HF (hydrofluoric) etching, or ion-beam etching.Precision Microfab, LLC (Arnold, Md.) is able to laser etchmicrochannels in a glass substrate. Micronit Microfluidics, BV(Enschede, The Netherlands) offers direct glass to glass bonding with noadhesive.

Other manufacturing processes and substrates can also be used in makingthe invention, e.g., polymer molding techniques can be used with apolystyrene substrate. Yet, compared to glass, polymer designs willtypically present added design challenges, particularly in relation tohydrophobicity. This topic is discussed under “Design Considerations”below.

The above-described embodiment will be shipped clean and sterile andwill typically be disposable (or kept by the patient's family as amemento) after a single use.

Numerous variations of the invention are possible, including highlyelaborate ones.

3. Fluidic Circuit Assembly (FCA)

The preferred embodiment not only satisfies the basis for a completeprenidial incubator system that combines easy access to a patient alongwith proper thermoregulation and fluidic ventilation at all times, butit is also simple enough that it can be manufactured in a way thatresults in an ultrapure environment for the patient such that chemicaland other forms of exposure (e.g., to electrical energy) are avoided.However, despite the inherent advantages of simplicity, it iscontemplated that elaborate embodiments of the invention may also bedesirable, particularly as science advances.

To satisfy this possibility, my invention further comprises a fluidiccircuit assembly (FCA). The FCA is not a distinct invention because itforms an integral part of the practice of elaborate embodiments of theinvention; in fact, the preferred embodiment may be taken to incorporatea simplistic version of the FCA. In order to manufacture highlyelaborate embodiments of the invention, it is necessary to employ anelaborate routing of microfluidic channels in intimate combination witha microcradle and any number of sophisticated devices. My inventionsatisfies this routing and integration requirement by employing an FCA.

A vented microcradle is an essential component of an FCA according tothe invention. Compared to the prior art, what is distinct about the FCAof the claimed invention is that is integrated with a vented microcradlefor the care of a prenidial infant.

By analogy, accomplishing an elaborate routing of microfluidic channelsis like a similar challenge solved in the field of electronics using aprinted circuit board (PCB), except in this case vias and traces arereplaced by vias and channels. The channels and vias are formed inlayers of “board” that are laminated together. The result is a fluidiccircuit board, or FCB. A variety of small-sized devices can be directlymounted to an FCB to form a fluidic circuit (board) assembly (FCA) in amanner similar to mounting devices on a PCB to form a printed circuit(board) assembly (PCA). Furthermore, a PCB/PCA may be included in an FCAto enable integrated electronic control and monitoring of a microfluidicsystem. Devices can be added or replaced by mounting them to the FCB atany time. This eliminates the need to rely on a single monolithicdevice, thus helping to speed development time, to linearize therelationship between cost and complexity, and to improve versatility andcustomization down the road.

The term “FCB” may also be used to mean an “FCA” in the same way thatthe term “PCB” is often used to mean a “PCA”. That is to say, an FCB mayrefer not only to a plain FCB but also to an FCB populated withcomponents, or more specifically an FCA.

An FCA is a type of integrated microfabrication technology (IMT) and canaccommodate other IMT technologies (i.e., submillimeter, micrometer(micron), submicron, and nanometer technologies). In addition to beingable to support a large variety of small-sized components (e.g.,microfluidic components, microelectronic components, microsensors,micro-electro-mechanical (MEMS) devices, labs-on-a-chip, etc.) the FCAcan also accommodate intimate combination with various large scaledevices (e.g., a microscope, thermal imaging camera, spectroscopydevices, chemical analysis equipment, etc.). According to the invention,the FCA may also include machine-readable indicia or markings as well asa temperature bath to maintain ambient temperature for the incubatorenvironment and various FCB components.

Referring to FIG. 5, a generalized embodiment of an FCA 400 according tothe invention comprises a plurality of wafer or board layers 24 (e.g.,made of glass) laminated together (e.g., using direct bonding of glassto glass without adhesive). The FCA 400 includes the following minimumfeatures: a network of microfluidic channels 25 patterned in boardlayers 24 to serve the cradle 26 of a vented microcradle with fluid.Microfluidic channels and vias patterned in an FCB may connect or serveany number of devices or components 27. The FIG. 2D embodimentincorporates a simplistic FCA involving two board or wafer layers, thelayers being formed by the glass petri dish 6 and the glass sheet 7,which are laminated together using direct bonding without adhesive;microfluidic channels 5 and via 10 are patterned in one of the boardlayers (glass sheet 7) to serve the cradle 1 of the vented microcradlewith fluid. In general, note that vented microcradles (and, hence, anFCA, which incorporates a vented microcradle) are easily distinguishedfrom prior art enclosures for prenidial infants because theyspecifically combine a cradle 1 having an open top 2 with a microfluidiccirculator system. A vented flooring according to the parent teaching(Ser. No. 10/908,861) may also be incorporated within the scope of anFCA.

FIGS. 6A-D illustrate embodiments of a side-vented microcradle employingFCAs of modest complexity limited to three board layers. Referring toFIG. 6A, a micropump 28 mounted to the bottom of an FCB 29 causes afluid incubation medium M to be urged through a network of microfluidicchannels 5 so as to fluidically ventilate a patient P inside theside-vented microcradle 1. Various devices (e.g., a flow meter, pHmeter, etc.) can be mounted to the FCB. Note that in contrast to theFIG. 2D embodiment, the embodiments shown in FIGS. 6A-6D enable a returnpath for fluid by adding additional ventilation ports 30 for thispurpose. In FIGS. 6A-B the additional ports operate vertically whereasin FIGS. 6C-D they operate horizontally. In FIGS. 6C-D the uppermostboard layer provides a retaining wall 31 for a microdrop μD under whichthe patient P is submerged. FIG. 6D illustrates a mounting of devices Dto both top and bottom sides of an FCB. Although fluid M is shown byarrows to be circulated in one direction, it is understood thatto-and-fro or reversible circulation can be accomplished with anappropriate pumping scheme. In FIG. 6D, dual or opposing pumps areprovided.

FIGS. 7A-B show an easy method of making the FIG. 6B embodiment usingglass sheets for the board layers 32-34. Referring to FIG. 7A,microfluidic channels 340 are etched on the bottom side of the topmostlayer 32; vias are etched through the layer 32, including vias 35 forfluidic communication with devices 36 mounted on top (shown in outline),vias 37 for ventilation ports 30, and a via 38 forming the top half ofthe cradle walls of the microcradle 1. Referring to FIG. 7B,microfluidic channels 39 are etched on the bottom side of the middlelayer 33; vias are etched through the layer 33, including a via 40 forfluidic communication with a device mounted on top of the FCB and a via41 forming the bottom half of the cradle walls of the microcradle 1. Thebottommost layer 34 does not require micromachining.

FCBs allow for versatile placement of component devices. Devices can bemounted on the top, sides, or bottom, or even inside the FCB. The FCBcan also support devices that are placed above a microcradle or thatreach into the microcradle. Referring to FIG. 8, a chamber 42 can beetched in a layer 43 to support a device 44 inside the FCB. Devices canalso be placed in microchannels or in the microcradle itself.

FCBs also allow for versatile placement of electrical connections.Conductive traces and vias can be directly patterned on the board layersof an FCB in a manner analogous to PCB manufacture. Referring to FIG.9A, since raised traces between layers would interfere with lamination,channels 45 can be etched in a board layer and traces 46 can then bedeposited in the channels so that the layers 47 will make flush contactwith each other. Referring to FIG. 9B, component leads 48 of a device 44mounted to an FCB are inserted through holes in a PCB in the manner ofthrough-hole construction and attached with metal solder 49. FCBs willgenerally be used only once for health reasons; yet it may still bedesirable to recover some of the components on an FCA for repeated use.Referring to FIG. 9C, component leads 48 are inserted into a solderlessconnector 50 for easy removal; a removable anchoring means 51 (e.g., ascrew or snap-in connector) can be used to mechanically fix a componentdevice 44 to the FCB; a sealing means 52 can be used to seal a fluidicconnection between the component device 44 and a microfluidic via 53 inthe FCB. Referring to FIG. 9D, a PCB can be mounted to an FCB andcomponent devices 44 can be mounted using surface-mount constructionsuch that component leads 54 are soldered to pads on the PCB; a ventedmicrocradle VM is shown in this figure; FIG. 9E shows a cross-sectionalview of this surface-mount arrangement. A PCB may be recessed into anFCB board layer by etching into the layer. Insulation can be added forelectrical isolation.

Devices requiring electrical support can be powered by a power supplylocated on or off of an FCB. Power can also be communicated wirelessly;for example, photoelectrical energy can be used to power a deviceembedded beneath transparent layers.

Some devices may require pneumatic support. Microfluidic channels cansupport both liquid and gaseous fluid flow. A network of gaseousmicrofluidic channels with a system of automated valve manifolds can beemployed to connect a plurality of pneumatically-driven devices on anFCB. A similar arrangement can be used with liquids to communicatehydraulic power to devices. Gaseous channels can be used for otherpurposes as well (e.g., to serve chemical processes on a lab-on-a-chip,to maintain pH in a fluidic reservoir employing a CO₂ buffering system,etc.).

Optical fibers (fiber optics) along with lenses may be routed orembedded inside an FCB or in devices supported by the FCB. Fiber opticsmay be employed for visualization, data transfer, and illumination.

An FCA may include reservoirs for fluid, e.g., for sequential media, tocondition the fluid incubation medium, to serve chemical processes on alab-on-a-chip, etc. Referring to FIG. 10, the FCB may support fluidsupply lines 55 and fluid microcontainers 56, including pressurizedlines and microcontainers for gas or liquid; the FCB may include varioustypes of male and female connectors 57 for fluid supply lines 55,microcontainers 56, and for filling or emptying reservoirs; the FCB maysupport wells 58 and microdrops 59 for liquid; the FCB may includeembedded reservoirs 60 for fluid. Microcradles and even microchannelsthemselves also provide reservoirs for fluid in an FCB. Droplets 61 maybe stored for digital microfluidics.

To connect an FCA externally, in addition to fluid supply lineconnectors, the FCA may also support connectors (or ports) forelectrical lines and optical fibers. The FCA may also support wirelesscommunication using both electronic and optical (e.g., infrared)signals. The FCA may also support mechanical connections for guide wiresor other mechanisms to actuate devices mechanically. FCAs support avariety of interconnects.

For operation in a fixed position, an FCA is mechanically fixed orlocked securely into position, preferably using a removable means, forexample, a screw-down or clamping means, or alternatively a slotted,seated, or cartridge-type means, including any needed connectors oralignment guides. An FCB socket connection such as a PC-slot typeelectrical connection or a solderless-type electrical connection maydouble as a means of mechanically fixing the FCB into position; variousother connections (e.g., fluid line connections) may also serve thispurpose; connections may be modified to better serve this purpose. Aretaining clip, lock, or other means may be employed to preventaccidental loosening of the FCA.

Sensors and alarms may be employed to indicate whether the FCB is fixedproperly into position and to detect various conditions such as whetherconnections are properly made. In general, an FCA may supportplug-and-play technology such that the FCA is recognized as a device bya computer running relevant software. The software can then be used toreport various alarm or status conditions associated with the FCA deviceas detected by sensors.

In addition to including fluid ports for ventilation, a ventedmicrocradle may also include fluid ports along with related microfluidicsystems to satisfy a variety of other purposes as well, e.g., for dosingmedications, dispensing, flushing/filling, hygiene, therapy,conditioning, analysis, transport, etc. Fluid ports may be reserved fordedicated purposes (e.g., ventilation only) or for dual or selectablepurposes (e.g., ventilation and medicine delivery). Fluid ports mayinclude various features (e.g., nozzles, a particular pattern ororientation, large sizing to speed fluid exchange, shutters or valves toprevent backflow or undesired mixing, etc.) to complement theirfunction. Similar diversity may be achieved with respect to fluid portssituated elsewhere on an FCA, e.g., at the site of a reservoir,lab-on-a-chip, etc.

The variety of analysis purposes satisfied by microfluidic systems hasbeen increasing; examples include compound screening and profiling,diagnostics and point-of-care (POC) testing, chemical analysis,proteomics, etc. Point-of-care testing means testing in proximity to apatient as opposed to sending a sample away to a lab for testing.

FCAs can support digital (discrete droplet) microfluidics as well asanalog (continuous flow) microfluidics.

FCAs can support programmable microfluidic networks, includingprogrammable valve arrays. FCBs can support removable covers to sealmicrofluidic vias and channels at places where optional or alternatecomponents can later be added or removed as desired.

FCAs can be populated with a great variety of components and can beintegrated with a great variety of larger scale equipment.

To give only a small sample of the possible variety, FCA components mayinclude microfluidic, electronic (e.g., electrical, microelectronic, andcomputing), magnetic, chemical, biochemical, physical, mechanical,physiological, electrophysiological, biological, medical, radiological,optical (e.g., fiber optic, photoelectronic, and optoelectronic), andacoustical devices and sensors; lighting and illumination devices (forboth visible and invisible, e.g., infrared, spectra); microsurgical,hygienic, and therapeutic devices; labs-on-a-chip, micro total analysissystems (μTAS), and micro-electro-mechanical (MEMS) devices;nanotechnology (e.g., coatings to make surfaces slipperier); fluidconditioning devices (e.g., for chemical and thermal conditioning offluid) and various environmental maintenance devices; patientconditioning devices (e.g., devices to apply coatings or treatments to apatient's body or eggshell); micromanipulators and instrumentpositioning devices; and patient handling or transfer devices. Basicmicrofluidic devices include pumps, valves, mixers, filters, injectionnozzles, and dosing devices. Notable sensors include sensors (andalarms) for pH, osmolarity, biochemicals or biochemical markers,temperature, heat output (microcalorimetry), pressure, internal pressure(e.g., a tonometer to measure hydrostatic pressure inside the egg),humidity, fluid reservoir levels, gas or liquid flow, biohazards (e.g.,pathogens, toxins, teratogens), chemicals, biochemicals, light,electricity, magnetism, viscosity, biological signals, electrophysiologydata, and data on patient growth or health status. Additionally, newdevices are constantly being developed.

To again give a brief example of the possible variety, FCAs may beintegrated with relatively large scale equipment such as equipment foroptical coherence tomography, nuclear magnetic resonance and magneticresonance imaging (MRI), thermal imaging, microscopy, spectroscopy, massspectrometry, electrophoresis, chromatography, fluorescence,electrochemical detection, chemical analysis, sonography,micromanipulation and instrument positioning, automation, computing,recording, communication, etc.

FCA designs are compatible with modular microfluidic architectures,which in turn may also be adapted to accommodate vented microcradledesigns. Referring to FIG. 5, an active component of a fluidicbreadboard (FBB) may be treated as a component device 27 with respect toan FCB containing a passive component designed for the FBB. According tothe FBB concept proposed by workers at the University of Illinois atUrbana-Champaign and Northwestern University, the FBB is conceptuallyand physically split up into two halves: one containing all of theactive components, and the other containing all of the passivecomponents. The active components include micro valves, pumps, mixers,sensors, heaters, and other elements that manipulate or sense fluids onthe FBB. The passive components consist of microfluidic channelnetworks. (Shaikh et al, “A Modular Microfluidic Architecture forIntegrated Biochemical Analysis,” Proceedings of the National Academy ofSciences of the United States of America, vol. 102, no. 28, pp. 9745-50,July 2005.) In general, FCB/FCA designs are more versatile than FBBdesigns because according to the FCA concept active components may beintegrated discretely instead of as a single, monolithic breadboardcomponent. Also, FCA designs can accommodate a plurality of FBB designs.Thus, the FCA is both breadboard and motherboard for a ventedmicrocradle, in the fluidic sense, to use a computer analogy.

Various modular microfluidic architectures are taught by Neukermans(U.S. Pat. No. 6,068,751), Kennedy (U.S. Pat. Nos. 6,086,740;6,488,895), O'Connor et al (U.S. Pat. Nos. 6,536,477; 6,729,352;6,827,095; 6,919,046), Bergh et al (U.S. Pat. Nos. 6,737,026; 6,749,814;6,890,493), Karp et al (U.S. Pat. Nos. 6,814,938; 6,880,576), Pamula etal (U.S. Pat. No. 6,911,132), and Zhou et al (U.S. Pat. No. 7,011,793).Zhou et al ('793) also teach a microfluidic breadboard.

In terms of its design architecture, an FCA according to the presentinvention may be construed as a specialized type of microfluidicmotherboard employing a vented microcradle for prenidial infant care.Microfluidic motherboards are known in the prior art. For example,workers at the Pacific Northwest National Laboratory employ amicrofluidic motherboard for bioanalytical applications comprising apolymer substrate containing a microchannel network and outlet fittings,along with various surface-mountable component devices (e.g., plug-inreservoirs, micropumps, microvalves, and sensors) populated on top ofthe substrate, and also including fluidic connection to externalequipment such as a mass spectrometer. (Lin et al, “Microfluidic Deviceson Polymer Substrates for Bioanalytical Applications,” In 3rdInternational Conference on Microreaction Technology; MicroreactionTechnology: Industrial Prospects, IMRET 3, Apr. 18-21, 1999 (Frankfurt,Germany), New York, N.Y.: Springer, pp. 451-60, 2000.) But unlike thepresent invention, prior art architectures do not include or serve avented microcradle for infant care.

As a side note, regarding a macroscopic fluid system, Laakaniemi et al(U.S. Pat. Nos. 4,110,140; 4,188,977) teach a fluid system circuitboard.

Those skilled in the art of microfabrication, and particularlymicrofluidics, will appreciate that a vented microcradle employing anFCA according to the invention can be practiced in a great variety ofconfigurations along with numerous relevant technologies.

4. Temperature Bath

According to my teaching in U.S. Pat. No. 6,694,175, a patient'stemperature must be monitored distinctly from the ambient temperature ofthe incubator environment, e.g., using a thermal imaging camera. Inkeeping with that teaching and that of the parent application (Ser. No.10/908,861), the invention accommodates and further provides a number ofcontrols to regulate patient temperature in response to patienttemperature readings. For in order to provide competentthermoregulation, the patient's temperature will need to be adjusted(e.g., due to activity or metabolism) based on patient temperaturereadings.

My present invention provides a temperature bath to maintain ambienttemperature for the incubator environment. The extent to which atemperature bath is needed depends on a variety of factors. For example,if thermal perturbations of the incubator environment are small andinfrequent then an air system in ambient contact with the incubator mayprovide sufficient thermal transfer by itself. However, a liquid systemflowing over the incubator will provide much more thermal transfer thanan ambient air system by itself, and so a liquid bath will buffer theincubator environment against larger perturbations in temperature.

The ambient temperature to which the patient is exposed regards thetemperature of the fluid incubation medium bathing the patient in themicrocradle. Alterations of the ambient temperature can be made in avariety of ways. The parent application (Ser. No. 10/908,861) teachesmixing media of fixed hot and cold temperatures to obtain a desiredtemperature or by heating or cooling the medium directly. The properlyheated medium is then entered into the microcradle environment toprovide the right ambient temperature. However, it is also desirable fora wider area of the environment to be at the proper ambient temperature,and not only the fluid medium that bathes the patient in the immediatesense.

A temperature bath according to the invention provides a means toregulate the ambient temperature throughout an FCA as a whole. Thetemperature bath employs a running fluid at a given temperature thatcirculates in thermal contact with the FCA to establish the ambienttemperature. The running fluid may be pumped or fed by gravity. Apumping means may be located on or off of the FCA. The temperature ofthe running fluid is set by a thermal reservoir (thermostaticbath/circulator), by mixing (merging) fluids of different temperatures,or by heating or cooling the fluid directly.

The running fluid can be water or another liquid or even gas. Preferredqualities of the running fluid include being non-hazardous and havinglow flow resistance, high thermal conductivity, and high specific heatcapacity. For a given substance, increasing the rate of flow of therunning fluid will increase thermal transfer. Adjusting the temperatureof the running fluid will enable the ambient temperature of theincubator environment to be set.

A desired temperature of the running fluid can be set and adjusted usingany variety of means. For example, heating or cooling the running fluidto obtain a desired temperature may be achieved by means of heating andcooling devices located on or off of the FCA. Also, a thermostaticbath/circulator provides a stable reservoir for fluid at a precisely settemperature. Lauda Dr. R. Wobser GMBH & Co. KG (Lauda-Konigshofen,Germany) makes thermostatic baths including a circulator pump, with bathtemperatures controlled to ±0.01 Celsius degrees. Hart Scientific, Inc.(American Fork, Utah) makes ultrastable thermostatic baths, with bathtemperatures controlled to ±0.001 Celsius degrees. The temperature of astream of fluid from a set-temperature reservoir can also be adjusted bymeans of heating and cooling devices to obtain a desired temperature.However, according to the invention, and as further described below, itis most preferable to mix streams of fluid from two or more reservoirsat different set temperatures to obtain a running fluid at a desiredtemperature.

As used herein, thermal pull-up and pull-down times refer to an amountof time taken to respectively raise or lower ambient temperature to adesired temperature. Slow pull-up/down times create a large lag time(i.e., pull-up/down time) between detection of a need to change ambienttemperature and completion of a response that establishes a new ambienttemperature. As Heidemann et al explain, “[C]ontrol performance islimited by the lag time between detection and response.” (p. 708,citation below.) Consequently, short pull-up/down times are needed tocontrol changes in ambient temperature as rapidly as possible.

Microscope stage warmers employing a resistive heating element are wellknown in the art of live cell microscopy. For example, Heidemann et alemploy a temperature-controlled aluminum ring in thermal contact with apetri dish to provide a microscope stage warmer for a cell culture; thealuminum ring is heated by thermal contact with resistive heatingelements. (Heidemann et al, “Open-Dish Incubator for Live Cell Imagingwith an Inverted Microscope,” BioTechniques, vol. 35, no. 4, pp.708-716, October 2003). However, in absence of a cooling means (e.g., athermoelectric cooler), resistive microscope stage warmers haverelatively slow pull-down times, being limited by heat dissipation intothe environment. Consequently, such microscope stage warmers do notenable a fine, rapid control of ambient temperature. Another problem isthat infrared radiation emitted by a resistive device in close proximityto a patient may be undesirable in a prenidial incubator withoutshielding; similarly, an electronic means of heating/cooling placed inclose proximity to the patient may introduce electrical effects that areproblematic for the patient or for sensing equipment.

Barsky et al (U.S. Pat. No. 5,119,467) teach a transparent film radiantheat source for use with incubators comprising an optically transparentcoating of indium tin oxide applied to the incubator walls. The coatingis electrically conductive and serves as a resistive heating element. Inthe art of in vivo microscopy the bottom of an observation dish issometimes coated with indium tin oxide to provide a heating source.

It is also well known to circulate a temperature bath around dishesplaced in a container to maintain a certain temperature with respect tofluid inside the dish (incubation medium). In this regard, Okolab(Naples, Italy) employs a Lauda temperature bath/circulator to pump arunning fluid at a preset temperature around petri dishes in a containerso as to provide a fluidic microscope stage warmer. However, thisapproach is limited in its ability to provide a rapid change in ambienttemperature (the temperature of the bath bathing the petri dishes)because the temperature bath/circulator (thermostatic reservoir) holds avolume of fluid and it will take a substantial amount of time for thebath to establish a new bath temperature that is equilibrated throughoutthe bath volume. This problem increases with the degree of temperatureprecision desired since the change in bath temperature must be slow inorder to avoid overshooting the desired temperature.

Another possible limitation of this approach is found when the runningfluid is allowed to circulate freely around the dishes; this limits therate of flow because too great a flow rate might cause splashing of thefreely flowing fluid. However, high flow rates may be desirable becausethey enable greater thermal transfer rates. Another problem is that afreely flowing fluid may take random paths as it circulates, making itharder to predict thermal transfer; in turn, this may result in anuneven temperature distribution. In contrast, housing the running fluidin channels or some kind of jacketing system would permit higher flowrates; also, controlling the path taken by the fluid would make iteasier to predict and control thermal transfer.

Water jacketing systems find diverse uses. Of particular relevance,workers at the University of California, Irvine employ a water jacketingsystem to provide a microfluidic flow cell with a constant-temperaturebath. (Goodrich et al, “Enzymatically Amplified Surface PlasmonResonance Imaging Method Using RNase H and RNA Microarrays for theUltrasensitive Detection of Nucleic Acids,” Analytical Chemistry, vol.76, no. 21, pp. 6173-8, November 2004.)

Enzelberger et al (U.S. Pat. No. 6,960,437) teach atemperature-controlled (glass) bed, such that a temperature bath flowsinside the bed to maintain a desired temperature for a microfluidic chipplaced on top and in thermal contact with the bed. They employ first andsecond temperature baths at different temperatures; initially, fluidflows from the first temperature bath through a first inlet into thebed, but then when the temperature of the bed is to be changed during atemperature cycle, fluid from the second temperature bath (at the nexttemperature in the cycle) is flowed into a second inlet, and the flow offluid from the first temperature bath is stopped. As fluid at the secondtemperature is flowed via the second inlet into the bed, fluid continuesto flow out of the bed via an outlet. (column 28, line 37 to column 29,line 5)

Enzelberger et al solve a different problem than solved by theinvention. According to their art, two discrete temperatures (e.g., 60degrees Celsius and 97 degrees Celsius) are required to promotedifferent stages of a chemical reaction, such as a nucleic acidamplification process. In contrast, according to the present invention,fine, rapid control is needed over a continuous range of temperatures.For human infant care in a prenidial incubator, the applicabletemperature range is centered about a temperature substantially near 37degrees Celsius.

There are a number of reasons why control is needed over a continuousrange of ambient temperatures in a prenidial incubator. For example, apatient may require a range of ambient temperatures over the course ofincubation; for example, if the patient's temperature is taken and istoo high, the ambient temperature may need to be lowered. In anothertype of example, sometimes it may also be necessary to relax (orperturb) a thermal condition in order to detect a condition of thepatient; for example, as is known to be the case in neonatal incubation,exercising thermal controls may mask a patient's condition; however, byrelaxing a thermal condition and monitoring the patient's progress to anew body temperature, valuable information can be gained about patientstatus and the influence of incubation parameters.

A temperature bath according to the invention overcomes the limitationsof the prior art and satisfies these needs by providing a continuoustemperature range with fast pull-up/down times and a fine control ofambient temperature. According to the invention, a desired temperaturefor a running fluid is established by mixing fluid from a plurality oftemperature baths using any combination of valves and mixers, either bycalculation or preferably with the aid of feedback controls from atemperature-detecting means. In principle, this is similar to mixing hotand cold water using a kitchen faucet to provide a desired temperature.To control ambient temperature, the running fluid can be made to flow atthe desired temperature through channels in an FCA, or through a waterjacket surrounding the FCA, or through a bed in thermal contact with theFCA.

The temperature bath may be implemented in a variety of ways. One way isto employ three thermostatic baths, consisting of a neutral bath and hotand cold baths. The temperature of the neutral bath is set substantiallynear 37 degrees Celsius. The other two thermostatic baths are set tomaximum (hot bath) and minimum (cold bath) temperatures. These maximumand minimum temperatures determine a range of ambient temperaturesprovided by the temperature bath. To achieve a temperature warmer thanthe neutral temperature, a stream of fluid from the neutral thermostaticbath is mixed with a stream of fluid from the hot thermostatic bath toobtain a running fluid at the desired temperature. To achieve atemperature lower than the neutral temperature, a stream of fluid fromthe neutral thermostatic bath is mixed with a stream of fluid from thecold thermostatic bath. Another way is to eliminate the neutral bath andmix streams of fluid from the hot and cold baths directly together.

Streams may be mixed in analog or digital fashion. In digital fashion, aselectable number of (nominally identical) streams at one temperatureare mixed with a selectable number of (nominally identical) streams atanother temperature by selecting a number of valves to open or closediscretely. In analog fashion, streams are mixed by selecting a numberof valves to open or close and controlling the proportion of theiropening; the opening proportion may be varied digitally or continuouslydepending on the valve and its means of actuation. A difference betweenanalog versus digital mixing of streams is that digital mixing requiresvalves that simply open or close, whereas analog mixing requiresadjustable valves. As an alternative to mixing streams of fluid from aplurality of thermostatic baths, a stream of fluid from the neutral bathmay be heated or cooled as it flows past heating and cooling devices toprovide a running fluid at a desired temperature.

As opposed to a stream, a running fluid may also be circulated in theform of droplets according to the art of digital microfluidics. Digitalmicrofluidics is an emerging branch of microfluidics employing a varietyof techniques to manipulate droplets of liquid as opposed to streams.For example, creating a difference in wettability between leading andtrailing edges of a droplet and a surface will cause the droplet tomove. Any combination of an electrical, optical, mechanical, magnetic,physical, or chemical means to create such a difference can be exploitedto cause a droplet to move; the difference in wettability is measured asa difference in surface contact angle between the leading and trailingedges of the droplet. Known techniques of this sort includeelectrowetting, opto-electrowetting, and optical wetting. In general,with regard to the temperature bath application of the presentinvention, there is concern about heating the running fluid as abyproduct of fluid manipulation; an extreme example is provided by adigital microfluidic technique relying on thermocapillarity in the formof a thermocapillary pump, which involves a differential heating ofleading versus trailing portions of a droplet within a microchannel.(Burns et al, “Microfabricated structures for integrated DNA analysis,”Proceedings of the National Academy of Sciences of the United States ofAmerica, vol. 93, no. 11, pp. 5556-61, May 1996; Pollack et al,“Electrowetting-Based Actuation of Liquid Droplets for MicrofluidicApplications”, Applied Physics Letters, vol. 77, no. 11, pp. 1725-6,September 2000; Rosario et al, “Lotus Effect Amplifies Light-InducedContact Angle Switching,” Journal of Physical Chemistry B, vol. 1108,no. 34, pp. 12640-2, 2004; Chiou et al, “Optical Actuation ofMicrofluidics Based on Opto-Electrowetting,” Technical Digest,Solid-State Sensor, Actuator and Microsystems Workshop, Hilton HeadIsland, S.C., pp. 269-72, June 2002.) In contrast to wettabilitytechniques, a technique being explored by researchers at the OpticalTrapping Group, University of St. Andrews uses optical tweezers tomanipulate very fine droplets using a focused laser beam (opticaltrapping). Fluid movement can also be created by means of manipulatingparticles or beads in a fluid that are subject to forces produced byvarious means (e.g., optical tweezers, magnetism, non-uniform electricfields, etc.), or by subjecting the droplet itself to such forces (e.g.,dielectrophoresis). (Calhoun et al, “Paramagnetic Particles and Mixingin Micro-Scale Flows,” Lab on a Chip, vol. 6, no. 2, pp. 247-57,February 2006; Zeng et al, “Principles of Droplet Electrohydrodynamicsfor Lab-on-a-Chip,” Lab on a Chip, vol. 4, no. 4, pp. 265-77, April2004.)

Digital microfluidics enables operations to be performed using digitalcontrols, often at high speeds, such as transporting droplets,dispensing droplets from a fluid stream or reservoir, splitting orcutting droplets, merging or mixing droplets, reacting droplets,analyzing droplets, and so on. Of note, circulation of fluid in discreteform (droplets) using digital microfluidics has been found to enablemuch higher flow rates than circulation of fluid in continuous formusing stream-based microfluidics. High circulation rates aid thermaltransfer. Droplets can be combined to form a stream or they can bestored to form a reservoir. Droplets at a desired temperature may bedispensed from a stream or reservoir bearing fluid at the desiredtemperature or by heating or cooling the droplets to obtain a desiredtemperature using a heating and cooling means; droplets from differenttemperature sources (i.e., neutral, hot, or cold sources) may be mergedto create droplets at a desired temperature. Droplets at the desiredtemperature can be circulated in the form of a running fluid such that adesired ambient temperature results from thermal transfer between thedroplets and the FCB/FCA or related devices, with assurances andfeedback controls provided using a temperature-detecting means.

The mixing (merging) of fluid streams (or droplets) at differenttemperatures to obtain a running fluid at a desired temperature ispreferably aided by means (e.g., valves and pumps, or, in the case ofdigital microfluidics, digital controls) controlled by feedback from atemperature-detecting means. An exemplary temperature-detecting meansmay include rapid-response thermistors or a thermal imaging system.Feedback from the thermal imaging system or other temperature-detectingmeans can be used to assess and control the impact of the temperaturebath provided by the running fluid on ambient temperature. The thermalimaging system preferably includes a microscope lens for infraredmicrothermography and may be the same system used to detect thepatient's body temperature.

Feedback controls enabled by a temperature-detecting means may include,for example, opening or closing valves to admit fluids of specifictemperatures to specific areas or devices on an FCA (e.g., to provideadded cooling to an area that is found to be overheated); controlling amerging of fluids at different temperatures to achieve a desiredtemperature; moderating a pump to increase a rate of fluid flow toprovide added thermal transfer; turning on a microheater to raisetemperature or turning on a microcooler to lower temperature; commencinga reaction on a lab-on-a-chip when proper temperature is achieved; etc.By using a layer material that is transparent to infrared radiation, athermal imaging system can detect temperatures throughout the FCA. Ingeneral, the thermal imaging system can also be used to checktemperatures indicated by other temperature-taking means associated withthe FCA (e.g., thermocouples, thermistors, pyroelectric sensors,thermopiles, and any other devices exhibiting temperature-dependentelectrical effects, and also nematic/thermochromic liquid crystalmicrothermography) to improve and verify their accuracy.

Calculations may also be used to predict and control a result of mergingfluids in terms of a resulting temperature achieved.

Assuming latent heats are not involved, then in general when two volumesV₁ and V₂ of the same fluid substance are mixed, the resultingtemperature T is given by the formula T=(V₁T₁+V₂T₂)/(V₁+V₂), where T₁and T₂ are the original temperatures. This formula can be converted toterms of relative proportion as T=(p₁T₁+T₂)/(p₁+1)=(T₁+p₂T₂)/(p₂+1),where p₁=V₁N₂ and p₂=V₂N₁. This reduces to T=(T₁+T₂)/2 when theproportions are equal. Correction for a variation in specific heatcapacity with temperature will seldom be necessary over a narrow rangeof temperatures centered about ambient temperature.

Correction will be necessary when a means of manipulating fluid volumescontributes heat. For example, if n droplets at T₁ are mixed with onedroplet at T₂, then the predicted temperature is T=(nT₁+T₂)/(n+1).However, if the process of manipulating the droplets adds heat, then thefinal temperature will be higher.

A temperature bath according to the invention, including separate ones,can be routed to devices on an FCA or to regions in an FCB havingdiverse thermal requirements, or even to external devices or equipment.In this way, ambient temperatures may be accurately maintained in aprenidial incubator by means of a temperature bath according to theinvention.

The prior art is crowded with efforts to ensure a general cooling ofelectronic circuits by means of a circulating fluid, with increasingemphasis being placed on a use of microfluidics. However, the presentinvention solves a more specific problem. Namely, the present inventionis concerned with establishing a specific ambient temperature.Nevertheless, of particular note is the teaching of Siebold et al (U.S.Pat. No. 5,142,441) for a circuit board containing an internal networkof microchannels for a running fluid to provide a cooling of devicesplaced thereon.

Other examples noted along these lines include the teaching of Daikokuet al (U.S. Pat. No. 6,351,384) for a device and method for coolingmulti-chip modules; Jun et al (U.S. Pat. No. 6,582,987) for a method offabricating a microchannel array structure embedded in a siliconsubstrate; Zeighami et al (U.S. Pat. No. 7,002,801) for a method ofcooling a semiconductor die using a microchannel thermosyphon; Nelson etal (U.S. Pat. No. 6,529,377) for an integrated cooling system in theform of a fluid-bearing tape; and, Goodson et al (U.S. Pat. No.6,991,024) for an electroosmotic microchannel cooling system. Also ofnote, workers at Duke University employ digital microfluidics(electrowetting-based techniques) to cool microelectronic circuits(Pamula et al, “Cooling of Integrated Circuits using Droplet-BasedMicrofluidics,” Proc. of the Association for Computing Machinery GreatLakes Symposium on VLSI, pp. 84-7, 2003.)

Those skilled in the art of regulating the temperatures ofmicrofabricated devices will appreciate that an ambient temperature fora vented microcradle can be accurately monitored and maintained using adiversity of means.

5. Machine-Readable Indicia

Bennett et al (U.S. Pat. No. 5,051,736) teach a method of determining alocation of an instrument over a glass surface of a computer screen bymeans of machine-readable etchings. At the time of their teaching ameans to produce such etchings was not readily available. However, todayion-beam etching provides a cost-effective means for producing extremelyfine etchings with remarkable detail. But the prior art does not teachor fairly suggest a use of machine-readable indicia or markings producedby ion-beam etching to aid in the automated control or positioning ofinstruments with respect to an incubator for premature infants or thepatient contained therein. Cecchi et al (U.S. Pat. No. 6,448,069) teacha use of indicia to identify members of a community of embryos in acompartmentalized structure. But because indicia are typically imprintedat the time of manufacture, a problem with the art of Cecchi et al isthat the means of identification is limited. To overcome thislimitation, the invention provides a means to superimpose text andimages on a display with the aid of a computer and software in referenceto indicia etched into the incubator in a machine-readable format at thetime of manufacture. The display can take the form of a hospital monitorfor doctors and nurses to look at.

According to the invention, machine-readable indicia etched into aprenidial incubator at the time of manufacture (e.g., on the surface anFCB layer) can be read into the computer and linked to informationstored in a data structure, for example, a patient's name or his or heruniquely assigned hospital identification number (Patient ID); in turn,for example, the information (e.g., the patient's name) can then besuperimposed onto the display for the benefit of health care personnel.For example, the patient's image, obtained by microscopy and captured bymeans of a digital (e.g., CCD) camera, can be displayed in a visualfield on the monitor along with superimposed information such as thePatient ID. Thus, by means of machine-readable indicia, the inventionenables personal and relatively unlimited information about a patient tobe displayed on a monitor and continuously updated in a user-friendlyformat.

Other exemplary uses of machine-readable indicia or markings are alsoachievable; for example, data concerning the model of incubatormanufactured can be etched onto a surface of the incubator and read intoa computer to tell the computer what software to run or what parametersto include when running the software. Similarly, rather than having tostruggle with space and size limitations to imprint indicia that arehumanly meaningful on the surface of a prenidial incubator, e.g., toidentify Medication Port 1, a machine-readable description can be etchedonto the surface and a humanly meaningful translation can besuperimposed on a visual display (monitor) by aid of a computer. Iconsand other user-friendly symbols (e.g., clickable symbols) can be shownon a display to indicate regions in a visual field wheremachine-readable indicia are embedded. Various information can be hiddenor displayed according to user preferences or in keeping with systematicevents. An optical means is preferred for reading the indicia ormarkings by computer.

According to the invention, with the aid of machine-readable indicia ormarkings etched onto a surface of a prenidial incubator, an instrument(e.g., a catheter or micropipette used in transferring a prenid) may bepositioned by a computer-controlled machine over an incubator enclosuresurrounding the infant (e.g., for the purpose of transfer). Additionalcontrols, both manual and computer-assisted, may also be implemented inthis scheme, e.g., with the additional aid of visualizing the infantoptically under the microscope. For example, micromanipulation andmicrosurgery may be implemented in this scheme. In general, a grid orother framework of machine-readable markings can be etched onto asurface of a prenidial incubator along with appropriate machine-readableindicia to enable automated control and positioning of instruments forpatient care and incubator maintenance.

According to the invention, artistic symbols, e.g., aesthetic orreligious symbols, can be etched onto a visible surface of a prenidialincubator for the benefit and comfort of viewers. For example, koalabears may be etched onto the surface of the flooring of the incubatorand matching bears may be printed on garments worn by nurses. Similarly,a dove might be etched in cases where parents plan to have their childbaptized in the incubator. An alternative to visible etching provided bythe invention is to simply have such symbols superimposed on a displayfor the benefit of viewers.

According to the invention, incubator units can be shipped with bar codelabels that match model and serial numbers imprinted on the incubator atthe time of manufacture. Other manufacturing data can also be included,e.g., for quality control purposes. Such data can be imprinted on theincubator in a machine-readable format using ion-beam etching so that acomputer can verify the correct correspondence, for example, to makesure that the incubator being used is of the proper type and that amix-up has not occurred.

Graduated markings (e.g., to indicate lengths, areas, degrees, etc.) maybe etched onto an incubator surface with units of measure indexed inmachine-readable form. Such graduations may be used to help measure thepatient's body size (morphometric) parameters, to help guide or positioninstruments, and so on. Markings may also be used to produce diffractiongratings or patterns and other optical effects. For example, adiffraction pattern can be used in the alignment or focusing of opticalequipment.

Ion-beam etching in a glass substrate is preferred for imprinting themachine-readable indicia or markings. However, other etching means orsubstrates may also be used.

Machine-readable indicia or markings may be etched anywhere on an FCAaccording to the invention. Binary encoding of indicia or markings ispreferred.

6. Transfer Catheter

FCB/FCA designs according to the invention are so versatile that theycan even assume instrumental forms, including the form of a catheter,cannula, aspiration needle, endoscope, laparoscope, or other instrument.A transfer catheter is an embodiment of the invention in a catheterform. The transfer catheter is used to transfer an infant from anincubator to the maternal body or, conversely, from the maternal body toan incubator.

Prenidial transfer means the transfer of embryos or hatchlings. In theprior art, hatchlings are seldom transferred because the prior art hashad no success in incubating prenids to the hatchling stage, and soprenids are typically transferred to the maternal body at the embryostage (i.e., before hatching). With respect to the maternal body,prenidial transfers may take place transcervically with placement in theuterine cavity or in a fallopian tube accessed via the uterine cavity;prenidial transfer may also occur laparoscopically. In a typicalscenario, after a period of external incubation, transfer to thematernal body will take place transcervically, with the infant beingplaced in the uterine cavity ready for implantation.

Referring to FIG. 11, to produce a transfer catheter according to theinvention, an FCB design is created in layers 62. Typically, the FCB iscut 63 to trim away excess board material, and the cut FCB 64 is shapedby any combination of forming, machining, and polishing to produce arounded catheter body 65. Microfabrication steps are generally includedin producing the transfer catheter. The transfer catheter may includeFCA devices/features located in/on the catheter body 65 and may becombined with external equipment.

Referring to FIG. 11, a Z-axis travels the catheter lengthwise, anX-axis is perpendicular to the laminated layers 62, and a Y-axis isparallel to the laminated layers 62. The prior art relies on extrusionto create a catheter body, resulting in isotropic flexibility in the X-Yplane. By employing laminated layers 62 to create the catheter body, thepresent invention is distinct from the prior art because it provides foranisotropic flexibility, such that the catheter is less flexible in theY direction than in the X direction. (In FCA designs, a Z-axis isdefined perpendicular to layers forming an X-Y plane; but an exceptionoccurs with respect to transfer catheters, in which case axes aredefined in view of compatibility with anatomical standards.)

The prior art generally teaches that a soft catheter is desirable toprotect the tissues of the mother's reproductive tract, but that thesoftness makes transcervical insertion difficult. The prior art facescompromise in this regard, generally employing either a harder catheteror a stiff sheath to aid in insertion. But the present invention offersan exceptional advantage in that a slight twisting motion willeffectively stiffen the catheter. Specifically, the catheter, when bowedin the X direction due to flexion caused by resistance to insertion,will tend to align along the Z-axis when twisted, so as to reduce thebowing, because the twisting rotates the more inflexible ZY-plane into aplane formerly occupied by the ZX-plane. Thus, with some art and tactilesensitivity on the part of an operator, using a slight twisting motionunder tension, including a back and forth twisting motion, the catheteraccording to the invention can be made to serve the purpose of greaterstiffness during insertion while retaining a greater softness overall.

Durometer measures the hardness of a material in terms of its resistanceto permanent indentation and is used to compare polymers, elastomers,and rubbers. High durometer generally equates to stiffness and lowdurometer to flexibility. Cecchi et al (U.S. Pat. No. 6,165,165) teachan embryo transfer catheter produced by extruding mixtures of resinswhich have different durometers, and varying the percentage of theresins in the mixture along the length of the catheter; consequently,the catheter has a varying stiffness from the distal end to an oppositeproximal end, the distal end being softer and the proximal end beingmore rigid; the result is a catheter that is non-abrasive at its distalend, and resistant to wobble at its proximal end. The present inventionrelies on a layer method, rather than an extrusion method along thelines of Cecchi et al. According to the present invention, layers 62 canbe provided that vary in durometer along the length of the Z-axisaccording to various polymer techniques; layers 62 of differentdurometer can also be interlocked or laced to provide differentstiffness qualities along the Z-axis; also, stiffening structures can beembedded in the layers 62.

The maternal body provides an infant incubator. And so in some sensetransfer implies transit from one incubator to another. This is also thecase when transfer is made between two manufactured incubators. In thepast, prenidial transfer has presented a discontinuity of incubation,because transfer catheters of the prior art do not qualify as incubatorsin themselves. Thus, an object of sophisticated transfer is to providefor a continuity of incubation.

In satisfaction of this object, the present invention enables aninstrument of transfer, such as a catheter, to serve as an incubator inits own right. Those skilled in the art of microfabrication willappreciate that this object can be accomplished with any desired levelof sophistication based on the FCB/FCA concept. Such instruments mayembody not only an incubator according to the invention but also otherfeatures as well, as are needed according to an overall function of theinstrument (e.g., microsurgical tools to operate on the maternal body,MEMS-actuated covers, valves, or shields to keep cervical mucus out,lights and fiber optics to visualize the maternal body or surroundings,etc.). Such instruments may also be practiced in combined form (e.g., acannula can be designed by fabricating a channel (lumen) along theZ-axis of an FCB that is large enough to accommodate a catheter).

Those skilled in the art of medical engineering will appreciate thatmodifications of the FCB/FCA concept can be analogously applied to avariety of arts, e.g., oocyte aspiration, so as to relieve a number oflimitations previously encountered.

A transfer catheter according to the invention may be construed as atransfer incubator combined with an instrument to effectuate transfer(e.g., the catheter body 65), which may be combined with a number ofauxiliary instruments. In other words, the transfer catheter may beconstrued as an instrument of transfer containing a transfer incubator.To achieve full sophistication, the transfer catheter preferablyincludes a number of desirable incubator features such as a ventedmicrocradle and a temperature bath or heating and cooling devices tomaintain ambient temperature for the patient to avoid thermal shock. Ameans of taking the patient's temperature according to my teaching inU.S. Pat. No. 6,694,175 is also desirable, such as the exemplary meansdescribed below for use in a transfer catheter.

Successful transfer to the maternal body is needed for patient survivalafter incubation in a prenidial incubator. A problem with prior arttransfer methods involves an amount of air and liquid fluid expelledfrom the catheter into the uterus during transfer. Although a generalconsensus is that a volume of fluid transferred should be kept to aminimum so as to avoid flushing the patient from the uterus ordisplacing the patient from a preferred site, a dilemma arises because alarger volume is also desirable to flush a patient who may otherwisestick to a wall of the catheter. Another problem is that the patient maybe drawn out of the uterine cavity or displaced from a preferred sitewhen the catheter is extracted.

Using a transparent model of the human uterus, workers at Tel-AvivSourasky Medical Center and Tel-Aviv University made simulated cathetertransfers to study different catheter loading arrangements. Regardingbackground on the problem, in a 2004 paper they report: “The technicalprotocol of embryo transfer (ET) has not been changed since it was firstintroduced three decades ago. Typically, the catheter is positionedwithin the uterine cavity with its tip 5-15 mm proximal to the fundus,whereupon its load is carefully injected and the catheter is thenextracted. [ . . . ]

Mechanical factors, such as uterine contractions, catheter type, themethod of loading the catheter, the placement of the catheter tip andphysician skills, have been proposed to explain some of the disparitybetween the embryonic development and pregnancy rates.” (p. 562,citations omitted.) FIG. 12 shows one of their typical catheter loadingarrangements. One of the conclusions of their study is particularlyinteresting: “The viscosity of the transferred liquid should be as closeas possible to that of the uterine fluid in order to avoid transport ofembryos towards the cervix.” (p. 562) (Eytan et al, “A Glance Into theUterus During In Vitro Simulation of Embryo Transfer,” HumanReproduction, vol. 19, no. 3, pp. 562-9, March 2004.)

Given such concerns, those skilled in the arts of microfluidics andmicrofabrication will appreciate that a transfer catheter according tothe invention presents unique alternatives to the traditional transferprotocol. For example, how could a transfer catheter be provided thatenables an embryo or hatchling to be flushed from a catheter into theuterus with an unlimited volume of fluid, without increasing an amountfluid in the uterus itself, and while employing a fluid with the sameviscosity as uterine fluid? Referring to FIG. 13, an embodiment of atransfer catheter according to invention offers a solution to thisproblem. A micropump 69 fabricated inside the distal end 66 of atransfer catheter 65 according to the invention circulates uterine fluid70, such that the fluid 70 passes in via inlets 71 and out via a ventedmicrocradle 72, so as to cause a prenid P to transit out of themicrocradle (transfer incubator) 72 and into the uterine cavity 73 toeffectuate transfer. The micropump 69 may preferably be a peristalticpump with pneumatic actuation. The pump 69 may be reversible to assistin loading the patient P into the transfer incubator 72. Because thefluid 70 being circulated is that contained by the uterine cavity 73itself, then unlike the prior art, any amount or volume of fluid 70 maybe circulated (e.g., for the purpose of flushing the patient out of thecatheter) without increasing the volume of fluid in the uterus;moreover, the fluid being circulated has the same viscosity as theuterine fluid 70 itself. Additionally, a circulation may be made tocontinue as the catheter 65 is being extracted, to prevent the prenid Pfrom being drawn out or displaced by a movement of the catheter 65.Thus, the present invention provides a means to overcome the prior artlimitations.

Those skilled in the arts of microfluidics and microfabrication willalso appreciate that a transfer catheter according to the inventionpresents other unique alternatives to the traditional transfer protocolas well. For example, according to the FCB/FCA concept of the invention,an instrument of transfer may be equipped with a mechanical means suchas a microgripper or other microdevice to mechanically move a patientfrom the instrument into a desired release location or position. As iswell-known in the art of microfabrication, microgrippers and relatedmicrofabricated devices (e.g., a microcage) are able to manipulate smallbodies and can be adapted to patient handling at the small size ofprenidial infants. (Ok et al, “Pneumatically Driven Microcage forMicro-Objects in Biological Liquid,” MEMS '99: Twelfth IEEEInternational Conference on Micro Electro Mechanical Systems. TechnicalDigest. Orlando, Fla., pp. 459-63, 1999.) In addition, microsurgicaltools can be included, including with the aid of visualization aids, toprepare a reception in the maternal body for the patient and also todeposit the patient with the aid of a securing agent or structure.

Those skilled in the arts of microfluidics and microfabrication willalso appreciate that a transfer catheter according to the inventionpresents an exceptional means of refinement even within the context oftraditional transfer protocols. For example, the invention enablesgaseous and liquid fluid samples to be urged through a catheter withgreater control. In the prior art, a syringe-catheter complex isemployed to urge fluids. However, according to the present invention, amicropump may be used instead. In such a case, there is no need for asyringe as with the prior art. With no need of a bothersome syringe,referring to FIG. 14, the proximal end of the catheter 65, which ishandled by the operator, may conclude with a housing 74 provided tocover internal devices and to allow for convenient handling by theoperator.

Referring to FIG. 14, means of wireless communication may also beincluded to establish communication between a catheter's devices andexternal equipment, whereby one or more functions of the catheter can bemonitored and controlled. Controls and indicators 75 may also beprovided on the transfer catheter 65 itself.

Although constraints will apply, thanks to the versatility of FCB/FCAdesigns a transfer catheter according to the invention can include manyof the features generally available to prenidial incubator design.However, certain features will still be peculiar to the needs oftransfer (e.g., articulating wires to flex the catheter, or echogenicstructures such as ridges to locate the catheter in the uterus underultrasound). To give an extreme example of versatility, a transfercatheter may even contain an incubator sophisticated enough to serve asa stand-alone incubator in its own right when docked with a supportsystem. However, in general it is contemplated that the transfercatheter will serve as an incubator only part-time (i.e., duringtransfer).

Referring to FIG. 11, the FCB design for the catheter is preferablycreated in polymer layers 62. However, a semiconductor, polysilicon,polydimethylsiloxane (PDMS), or other substrate for the microfabricationof active and passive components may also be employed as one or more ofthe layers 62, and a surface 68 of the catheter 65 may be coated with aprotective layer if necessary. The FCB generally includes a network ofmicrofluidic channels and vias and provision for a vented microcradle;however, a single lumen may suffice for delivery of a prenidial infant.Optical fibers and lenses, articulating wires and anchors, electricalcontacts, and sensors and devices may be embedded inside the FCB.Relatively large channels (lumens) may also be included for delivery ofsubstances and other purposes. To provide for clearer ultrasonicguidance, the catheter may include echogenic structures to disperseultrasonic waves according to the art. The distal end 66 of the catheterwherein the vented microcradle is located may be formed to a desiredshape. The proximal end 67 may be connected to external equipment,including fluid lines, and electrical, mechanical, and opticalequipment. Articulating wires routed and anchored inside the cathetermay cause the catheter to flex to a desired shape and angle, and thesewires may be actuated externally (e.g., via articulating levers) orinternally (e.g., via micromechanical actuation). The catheter may alsobe sheathed in a laparoscopic device or other housing. Components may bepopulated on the FCB surface at the proximal end 67, which does notnecessarily need to be rounded like the distal end 66.

The transfer catheter may be molded or formed into a fixed bent shape.As an alternative to articulating wires, MEMS-actuated devices may causethe catheter to flex. The catheter should have markings or anorientation structure at the proximal end 67 to define X-Y axes and atop, bottom, and sides to help guide the operator. The catheter may alsohave sensors to sense orientation. A microcradle may be housed at thetip or on the sides at the distal end 66. A proximal portion of thecatheter may have graduated markings to enable a depth of insertion tobe measured by the operator. A microcradle array may be housed by thecatheter in cases where fraternal twins are being transferred andindividual transfer incubators are preferred. A stream of fluid may bedirected along a surface of a sidewall of the microcradle to helpprevent the patient from sticking to the wall; coatings, structures, andmechanical means may also be used for this purpose.

The portion of the catheter at the distal end 66 where the infant ishoused is preferably made of a translucent material to aid inmicroscopic examination, e.g., to verify that transfer has taken place.However, optical fibers provided for illumination and visualization mayalso be used for this purpose. Optical fibers contained by the FCB maybe used to visualize the infant and tissues of the maternal body. Fiberoptics may also be employed in conjunction with the FCB to visualizeinstrument activities. Patient temperature can be detected via anoptical fiber by maintaining a temperature sensor in an optical field ofview, such that the temperature of the patient (along with other bodiesas well) can be determined by reference. The sensor provides a referencepoint within the field of view. Employing a calibration reference in aradiometer's field of view is discussed further under “DesignConsiderations” below.

FCB designs can be made of diverse materials. For example, surgicalmetal layers can be laminated in high vacuum after cleaning withhydrogen ion or they can be bonded anodically. Those skilled in the artof microfabrication, with particular emphasis on microfluidics, willappreciate that employing laminated FCB layers according to theinvention offers greater versatility, functionality, and miniaturizationin a transfer catheter design than the prior art, which relies onextrusion rather than lamination to form a catheter body and relatedlumens. As with FCA designs in general, a transfer catheter according tothe invention can be operated using diverse external equipment as an aidto operation.

The prior art does not teach or fairly suggest a transfer cathetermanufactured from laminated layers. Nevertheless, the following priorart references are of additional note. Wallace (U.S. Pat. No. 4,863,423)teaches a catheter and cannula assembly, such that when “an obstructionor tortuous path is encountered in the cervical canal . . . the cathetermay be gently rotated within the cannula to assist its advance” (column2, lines 49-52); he also teaches graduated markings on a proximalportion of the catheter and cannula to enable an operator to measureinsertion depth. Tao (U.S. Pat. No. 6,610,005) teaches a catheter systemutilizing a protective catheter sleeve for introducing a catheter intothe uterus without mucus contamination of an inner catheter. Bosley etal (U.S. Pat. No. 6,527,752) teach a transfer catheter that includesultrasonically reflective components or features to enhance itsvisibility under transabdominal or transvaginal ultrasound guidance.Thompson (U.S. Pat. No. 6,010,448) teaches a transfer catheter having alumen to deliver adhesive to aid implantation and fiber optics to aidsite selection. Bacich (U.S. Pat. No. 5,472,419) teaches a transfercatheter employing a special shape of a distal opening for transfertranscervically through the uterus into a fallopian tube ortranscervically into the uterus. Kamrava et al (U.S. Pat. Nos.6,758,806; 7,033,314) teach articulating wires coupled to anchors (dumbbells) to provide for articulation of a distal portion of a hysteroscopeusing an articulating lever; they also teach a hysteroscope having fiberoptics to aid in visualization and illumination of the uterus.

7. Design Considerations

Referring to FIG. 1, fluid flowing out through a ventilation port 4 istranslated from a horizontal direction into a vertical direction so asto flow past the patient P and out of the open top 2 of the side-ventedmicrocradle 1. Correspondence between the rate of fluid flow through theside vents 4 and a desired flow rate past the patient P can bedetermined by calculation. In general, the cross-sectional area of avertical cross-section of the microcradle 1 (A_(cradle)) times the rateof flow in the vertical direction (v_(vert)) equals a sum over allventilation ports of a cross-sectional area of the ventilation port(A_(vent)) times the rate of flow out of the ventilation port(v_(horiz)). Here, the rate (or velocity) of fluid flow is measured inunits of distance per unit time. See FIGS. 15-16. However, in operation,in the equation of FIG. 16, the cross-sectional area of the microcradle1 (A_(cradle)) will actually be replaced by an effective cross-sectionalarea, which may change along the vertical direction. The effectivecross-sectional area will take into account an amount of space taken upby the patient and any objects made present inside the microcradle 1. Ifall ventilation ports have the same cross-sectional area and rate offlow, then a ratio of vertical and horizontal velocities will observe aconstant proportion, determined by the cross-sectional area of themicrocradle (A_(cradle)) and the net cross-sectional area presented by asum of all ventilation ports (A_(ports)), such thatV_(vert)/V_(horiz)=A_(ports)/A_(cradle).

Flow through the ventilation ports 4 may observe a directional patternso as to cause any number of desired flow effects in the microcradle(e.g., vortex flow). Flow out of a ventilation port may also bedeflected to cause a desired effect (e.g., deflection in an upwarddirection upon exit may be caused by a structure impeding a path offlow). In this way, a desired circulation can be achieved and problemslike having areas of stagnation can be avoided.

Drawn in this case more closely to a true relative proportion offeatures, FIG. 17 shows an exemplary mask pattern for the microfluidicchannels 5 of a side-vented microcradle along the lines of FIG. 2C. TheFIG. 17 design is based on a three-stage multiplexer (multi-splitter). Amultiplexer design such as this ensures that a path length of fluidtraveling to each ventilation port 4 is the same and that flow isbalanced among the paths. This design employs constant-velocity stages,such that a net sum of the cross-sectional areas of the channels in eachstage is conserved by starting with a wide channel (the 0^(th) stage)and then narrowing channels for each stage in succession (stages 1-3);alternatively, successive stages can be etched at different depths toconserve net cross-sectional area and, hence, a constant rate (velocity)of fluid flow. In another example, FIG. 18 shows a design which, likethe FIG. 17 design, ensures a constant path length for fluid travelingto each of the ventilation ports 4. But unlike the FIG. 17 design, thisdesign employs non-constant velocity stages; because the microchannels 5maintain the same cross-sectional area, velocity (v) in the channelsdecreases as the channels are split successively into more channels.

In general, it is desirable to conserve path length among ventilationports because this ensures fluid will arrive at each of the ports at thesame time. However, FIG. 19 shows a design in which path length is notconserved. This design serves as a substitute for the FIG. 6Bembodiment, such that only two glass layers are required instead ofthree. In this case, microchannels and vias are etched in a top layer 76and then bonded to a bottom layer; referring to FIG. 6B and FIGS. 7A-Bfor comparison, the top layer 76 of the FIG. 19 embodiment replaces topand middle layers (32, 33) in the FIG. 6B embodiment. A via 77 forms thewalls of the vented microcradle 1 that were formed together by vias 38,41 in FIGS. 7A-B.

Referring to FIG. 19, devices 36 can be mounted underneath the bottomlayer by etching vias (35, 40) in the bottom layer rather than in thetop layer 76; similarly, devices can be mounted on the surface of bothtop and bottom layers. Devices 36 can be arranged, for example, in ageometry that reduces the path length of channels (e.g., in acircumference around the microcradle) or, for example, in a manner thatleaves a greater distance between a device 36 and the patient.Non-limiting design considerations such as these illustrate versatility.

The fluid incubation medium is an aqueous physiologic solution. Water isattracted to a hydrophilic surface and ‘repelled’ by a hydrophobicsurface. Glass is hydrophilic and so water will automatically fillchannels and vias etched in glass based on capillarity. In contrast,many polymers are hydrophobic or not as wettable as glass, and sofilling channels and vias with an aqueous solution introduces a numberof design considerations; otherwise, channels and vias may not fill,resulting in device malfunction.

Whether a material is hydrophobic or hydrophilic is not so importantonce channels and vias have actually been filled. As McNeely et al (U.S.Pat. No. 6,296,020) explain, “[O]nce water has been introduced into the[capillary] tube the flow rates of the water are dependent more onpressure gradients and friction and less on whether the material ishydrophobic or hydrophilic.” Yet getting fluid to fill channels and viashaving non-wettable surfaces can be problematic. One approach is simplyto alter the surface of a polymer to make it hydrophilic after channelsand vias have been formed; coatings, UV exposure, and oxygen plasmaexposure are commonly employed. But these methods may introduce unwantedchemical considerations; also, a shelf life is introduced in cases wheregiven treatments deteriorate over time.

Another approach, which may also include altering the wettability ofspecific regions with treatments or coatings, is to engineer channels toensure a balanced urging of fluid, including through split (or forked)channels, without having to pipette forked branches individually.Without engineering of this sort, often what happens is that one half ofa forked branch will fill with water and the other will not, since thepolymer channels are hydrophobic. But with proper engineering fluidurged through a given channel can be made to fill tributaries equally.Similar considerations apply to combining and mixing fluid streams.

Examples of such engineering considerations include teachings commonlyassigned to BioMicro Systems, Inc. (Salt Lake City, Utah), including theteachings of McNeely et al (U.S. Pat. Nos. 6,296,020; 6,591,852;6,601,613; 6,615,856) and Lei et al (U.S. Pat. No. 6,637,463); theteachings of Jeon et al (U.S. Pat. Nos. 6,705,357; 6,883,559); as wellas teachings commonly assigned to Nanostream, Inc. (Pasadena, Calif.),including the teachings of Pezzuto et al (U.S. Pat. Nos. 6,418,968;6,748,978), Dantsker et al (U.S. Pat. No. 6,499,499), O'Connor et al(U.S. Pat. Nos. 6,755,211; 6,919,046; 6,981,522), and Karp et al (U.S.Pat. Nos. 6,845,787; 6,877,892; 6,890,093; 6,935,772). McNeely et al('020) also teach a use of air escape channels to facilitate complicatedfluid processing.

Also of particular note, Bhullar et al (U.S. Pat. No. 6,451,264) teach afluid flow control in curved capillary channels; Fuhr et al (U.S. Pat.No. 6,727,451) and Karp et al (U.S. Pat. No. 6,880,576) teach amicrofluidic control of microparticles (e.g., solid catalyst media)suspended in channels; Griffiths et al (U.S. Pat. No. 6,733,730) teach amethod and apparatus for reducing sample dispersion in turns andjunctions of microchannel systems; Weigl et al (U.S. Pat. No. 6,743,399)teach a pumpless microfluidic device that is entirely driven by areadily available force, such as gravity, capillary action, absorptionin porous materials, chemically induced pressures or vacuums (e.g., by areaction of water with a drying agent), or by vacuum and pressuregenerated by simple manual action, rather than by an external fluidicdriver requiring a separate power source having moving parts; Ismagilovet al (U.S. Pat. No. 6,843,262) teach fluidic switches and methods forcontrolling flow in fluidic systems; Lim et al (U.S. Pat. No. 6,866,067)teach a micro channel unit having a shape designed to reduce a pressuredrop when fluid passes through a connecting channel portion; Gilbert etal (U.S. Pat. No. 6,883,957) teach an on chip dilution system; and, Kimet al (U.S. Pat. No. 6,901,963) teach a microfluidic device forcontrolling a flow time of fluid.

As the above references make clear, the available body of knowledge toassist in making design considerations is growing rapidly in the arts ofmicrofluidics and microfabrication. By keeping up-to-date in the art,one of ordinary skill will be able to make obvious modifications to thepresent invention on a continuing basis. Also, many inventivetechnologies will be devised over time to complement and improve thepractice of the invention. The science of the invention is highlyinterdisciplinary and draws on many fields, and not only in the “micro”dimension, but also in larger and smaller dimensions as well.

Turning to the subject of visualizing the patient, Campbell et al (USpublished application 2002/0068358) disclose a window with an accessorylens for visualizing an embryo contained in a microfabricated chamber.Thompson et al (U.S. Pat. No. 6,673,008) teach windows along with meansof fiber optics for visualizing an embryo in a chamber.

Workers at Stanford Medical School's Network for Translational Researchin Optical Imaging are developing a MEMS-based dual-axes confocalmicroscope. (Wang et al, “Dual-Axis Confocal Microscope forHigh-Resolution In Vivo Imaging,” Optical Letters, vol. 28, no. 6, pp.414-6, March 2003.) Small in size, MEMS-based microscopes such as thiscould be developed for introduction like a probe into the fluidincubation medium of a prenidial incubator to visualize the patient. Asopposed to a probe-type MEMS-based microscope, a MEMS-based microscopemight also be designed for physical integration with the prenidialincubator itself, as a structural part of the microcradle or inside orin close proximity to it.

A vented microcradle employing a vented flooring according to the parentapplication (Ser. No. 10/908,861) offers a number of advantages. But anotable limitation overcome by the present invention is anincompatibility with a number of important microscopy techniques.However, if such techniques were replaced by a MEMS-based technology,the limitation would vanish. Essentially, the problem is that the ventedflooring may interfere with an optical path needed for a number oftechniques. These techniques minimally require a clear view from belowthe patient, as is required by an inverted microscope, or more generallya clear optical path in a vertical direction from above and below thepatient. A MEMS-based solution would obviate this requirement byenabling an optical path to be established laterally or with a probe.However, in absence of a MEMS-based solution, a side-vented microcradlewith a clear bottom and an open top offers the advantage of fluidicventilation combined with a clear vertical path for optics.

Referring to FIG. 20A, a temperature-detecting infrared camera 770(e.g., a quantum well infrared photodetector (QWIP) camera) is attachedfrom above to a microscope lens 78 for purposes of infraredmicrothermography to detect the body temperature of a patient P inside aprenidial incubator 79 (U.S. Pat. No. 6,694,175). In this case, it doesnot matter if the floor 80 of the incubator is opaque because theinfrared radiation being detected is emitted light. The same arrangementcan be achieved with focus from underneath provided that the flooring 80does not block or distort the infrared radiation. Referring to FIG. 20B,a microscope 81 is positioned with its objective (lens) 82 over thepatient P. In this case, light from a light source 83 beneath theflooring 80 will provide illumination for many ordinary viewing purposesprovided the flooring 80 is transparent enough. Light shining from abovemay also provide illumination; the illumination can be enhanced if theflooring 80 or other structures (e.g., walls) are mirrored orreflective.

If the flooring 80 has a pattern of vents according to the parentteaching (Ser. No. 10/908,861), then making the vented flooring 80 outof a material with a refractive index more closely matched to that ofthe physiologic solution (incubation medium) filling the vents will helpto reduce a distortion of light passing through the vented flooring 80;digital corrections for distortions can also be made by computer.Techniques like these may make it possible to clearly view the patient Peven through the vented flooring from below (i.e., using an invertedmicroscope).

Referring to FIG. 20B, some microscope techniques require light withspecial qualities, and so filters, polarizers, compensators, and otheroptical means (not shown) may be placed in the optical path.Consequently, even when viewed from above, a vented flooring 80 maystill detract from a desired quality of light by causing distortions oflight quality. These distortions may be reduced to some extent either bymaking digital corrections or by controlling properties of the ventedflooring 80, such as refractive index, diffraction, birefringence, grillpattern, and so on. Nonetheless, full compatibility with a number oftoday's high end microscopy techniques will generally require a clear,undistorted optical path as provided by a side-vented microcradle with aclear bottom and an open top according to the present invention.

Accordingly, when certain high end microscopy techniques are necessary,and assuming an optical MEMS implementation is not available, then aside-vented microcradle with a clear bottom and an open top according tothe invention will likely provide the best choice. However, a number ofsuch techniques introduce strict design requirements. Those skilled inthe arts of engineering and microscopy will appreciate a need to observea number of design considerations in this regard.

FIG. 20C illustrates a familiar setup for high resolution microscopyusing an inverted microscope having a high numerical aperture objectivelens 84. Light from a light source passes through a condenser 85 toilluminate a biological specimen S in a petri dish 86. The specimen Smay be covered with various mounting media, but for in vivo microscopyapplications the medium is a physiologic solution MM. Typically, thespecimen S rests or is plated on a coverslip (cover glass) CG that hasbeen affixed using adhesive 87 over a hole in the bottom of the petridish 86. A working distance WD for an objective lens 84 is defined asthe distance between the objective front lens element LE and the bottomof the cover glass CG above.

High numerical aperture objective lenses 84 are designed with aparticular immersion fluid IF in mind. The importance of the immersionfluid IF is its refractive index. The immersion fluid fills the workingdistance WD space, meaning the space between the objective front lenselement LE and the bottom of the cover glass CG above. Air (dryimmersion), water, and oil are the most common immersion fluids IF;glycerol/water mixtures are also used. Dry immersion objectives offerlarge working distances up to ˜2 mm, whereas oil immersion objectivesand water immersion objectives generally rely on relatively shortworking distances, for example, on the order of 220 microns (0.22 mm).

According to theory, best optical performance is achieved when themounting medium MM, specimen S, cover glass CG, immersion fluid IF, andthe objective front lens element LE all have the same refractive index;however, if imaging is limited to a focal plane immediately adjacent tothe cover glass CG, e.g., to a surface region of cells making directcontact with the cover glass CG, then only the refractive indices of thecover glass CG, immersion fluid IF, and objective front lens element LEare important. With present technology, objective lenses 84 are made ofglass having a refractive index of 1.515; consequently, oil immersionobjectives offer the highest numerical aperture (a measure of resolvingpower) because some oils have the same refractive index. However,cellular matter has an index of refraction ranging from about 1.33 to1.39 and a physiologic solution has a refractive index of about 1.33.Thus, when imaging beyond a focal plane immediately adjacent to thecover glass CG, some refractive index mismatch is inevitable. As aconsequence, with present technology, water immersion objectives allowfor imaging of the highest resolution through cells or aqueous to adepth of 200 microns past the cover glass CG; in contrast, oil immersionobjectives offer higher resolution for imaging limited to a focal planestrictly adjacent to the cover glass CG, such that focus through anaqueous layer is avoided. In general, high resolution microscopytechniques can be particularly sensitive to a thickness and index ofrefraction of the cover glass CG. The thickness of the cover glass CGmay nominally be 170 microns with an index of refraction of 1.515; othervalues are possible. The objective 84 may also include a correctioncollar to compensate for cover glasses with non-ideal values. Coverglasses may also be selected based on other properties such astransmission to certain wavelengths of light, polarization,birefringence, florescence, and so on. See Nikon's MicroscopyU website(www.microscopyu.com) for an in-depth discussion.

Referring to FIG. 1, the present invention generally enables highresolution microscopy using an inverted microscope by specificallyemploying a layer for the clear bottom 3 of a vented microcradle havingdesirable optical properties such as thickness, refractive index, and soon. Using FIG. 6B as an example, to enable high resolution microscopy inconjunction with an embodiment of the invention by employing an invertedmicroscope with a high numerical aperture objective, such as a Nikon(Melville, N.Y.) 60× plan apochromat correction water immersionobjective featuring a 1.2 numerical aperture and a 220 micron workingdistance, the bottom layer 34 is preferably a cover glass having athickness of 170 microns and a refractive index of 1.515. Othermodifications of the invention to enable high resolution microscopy willbe appreciated by one skilled in the art.

Micronit Microfluidics, BV (Enschede, The Netherlands) offers amicrofluidic chip having a 145 micron thick cover glass bottom to enableviewing by means of confocal microscopy inside a microchannel etched inthe bottom of an adjacent upper layer.

Referring to FIGS. 3-4, an embodiment of a temperature bath according tothe invention employing a double-walled vessel 16 would generally not besuitable for use with high resolution microscopy if, as shown in FIG. 3,an optical path 88 though the clear bottom 3 encounters the runningfluid 19 and an additional layer of glass 89. In other words, as shownin FIG. 4, even though the clear bottom 3 remains “clear”, in general itwill not be optically suitable for meeting the strict requirements of anumber of high resolution microscopy techniques. Thus, to ensure optimumcompatibility with such techniques, it is preferable to route therunning fluid 19 around a clear area (forming an optical window) 90 in aline-of-sight with the optical path 88. Though maintaining laminar flowin the clear area 90 may improve clarity, for optimum compatibility therunning fluid 19 must be routed around the optical path 88.

Referring to FIGS. 21A-B, a variety of temperature bath according to theinvention is provided in the form of a temperature-controlled (glass)bed 91, which preferably takes the form of a microscope stage, such thatthe temperature bath flows inside the bed in the form of a running fluid19 to maintain an ambient temperature for an FCA (not shown) that restsin thermal contact with a top layer 92 of the bed 91. The thermalcontact may be aided by means of a heat sink compound thinly applied tothe top layer 92 or by means of pressing and holding the FCA against thetop layer 92. Though not shown, it is understood in practice that aclear bottom of a vented microcradle contained by the FCA is positionedover an open area 93 in the bed 91 to provide a clear area 90 for anoptical path for microscopy. The temperature-controlled bed 91 accordingto the invention is distinct from the temperature-controlled bed ofEnzelberger et al because it provides for a continuous range oftemperatures (as opposed to two temperatures) and an opening 93 toensure an area 90 for a clear optical path for microscopy.

Alternatively, referring to FIG. 22, which is likened to FIG. 21A, anFCA layer providing a clear bottom 3 for a vented microcradle (e.g., thebottom layer 34 of the FIG. 6B embodiment) can replace the top layer 92of the temperature-controlled bed shown in FIG. 21A to directly bond theFCA to a bed-like temperature bath 94. In FIG. 22, upper layers of theFCA (e.g., layers 32-33 in the FIG. 6B embodiment) are not shown.

In general, microfluidic channels and vias can be routed within an FCAwith any degree of complexity. This includes routing a running fluid fora temperature bath through an FCA in such a way as to leave an area fora clear optical path for microscopic viewing into a vented microcradlecontained by the FCA. In the art of electronics, traces and vias can berouted on a PCB with the aid of auto-routing software programs.Analogous skills applied to FCA channels and vias will be appreciated bythose skilled in the art of microfluidics. To give a non-limitingexample, referring to FIG. 23, a running fluid 19 is routed through anFCA in such a way that a clear area 90 is left to provide for a clearoptical path for microscopy. This example is similar to the FIG. 6Cembodiment.

For exemplary purposes, FIGS. 24A-C depict a variation of the embodimentshown in FIGS. 6B & 7A-B. Referring to FIG. 24A, note that a cover glassportion 95 of the bottom layer 34 may be pieced in instead of beingcontinuous. A design consideration being explored in this example isthat with present technology it is possible to directly bond two orpossibly three layers of glass without adhesive; however, formicrofluidic designs it remains challenging to directly bond more thanthree glass layers. Therefore, as long as this limitation persists, itis desirable to limit to three or less the number of FCA layers that arefluidically contiguous with a vented microcradle, since such layers needto be bonded without adhesive in order to help assure an ultrapureenvironment for patient care. In this sense, what fluidically contiguousmeans is that if the layers were to employ an adhesive instead of beingdirectly bonded, then the adhesive might leak through a seam or boundaryinto fluid that a patient will be exposed to in the vented microcradle.However, non-fluidically contiguous layers (or regions) do not presentthis problem, and so adhesive may be used to bond layers withoutcompromising the purity of the care environment. Thus, it may bedesirable to piece in a layer for a fluidically contiguous region sothat adhesive can be used to bond added layers in a non-contiguousregion.

For example, referring to FIG. 24A, layer regions 95 (e.g., the coverglass) and 96 can be directly bonded to layer 33, whereas layers 96 and97, including an optional intermediary layer 98, can be bonded to eachother with adhesive. FIG. 24A is a side cross-sectional view taken alonga line 99 with respect to FIGS. 24B-C. Note that channels must be routedto prevent a leakage of fluid along a vertical seam 100 where apieced-in layer 95 meets an adjacent layer 96. In this example, vias101-102 route a channel 103 from a lower layer 33 up into an upper layer32 and then back down to avoid the seam 100. In general channels andvias bearing potentially contaminated fluid (e.g., contaminated withadhesive residues) can be routed anywhere in an FCA provided that thechannels and vias are not contiguous with seams or boundaries that mightpermit contamination of the care environment.

Referring to FIG. 25 in comparison with FIG. 24A, routing of amicrofluidic channel 103 into an upper layer and back down again toavoid a seam would not be necessary if a bottom layer 104 werecontinuous (though of non-uniform thickness) rather than beingpieced-in, so that in effect layer regions 95 and 96 (having differentthicknesses) were formed by a single piece of glass. This approach wouldenable other layers (e.g., 97-98) to be added on with adhesive withoutcontaminating the care environment, while also providing a flat bottomfor a FCA. However, direct bonding of a glass layer 104 of non-uniformthickness as shown in FIG. 25 might be too challenging to perform withpresent technology due to a risk of cracking the glass. Yet etchingcould be performed after direct bonding of a layer of uniform thickness,resulting in a layer 104 of non-uniform thickness as shown in FIG. 25.But if a flat bottom for the FCA is not necessary, then something alongthe lines of FIG. 22 might be preferable.

Referring to FIG. 26, which is similar to FIG. 25, another option is todirectly bond layers 32-33, 105 to each other without any adhesive firstand to then bond an additional layer 106 with adhesive to layer 105. Forexample, glass layers 105-106 may nominally be 30 and 140 microns thickrespectively and have a refractive index of 1.515. Accordingly, thebottom layer 34 will total 170 microns in thickness and have arefractive index of 1.515 in the clear area 90 provided for an opticalpath, as is desirable for high resolution microscopy techniques. Greatflexibility is afforded by this approach. For example, layers 97 and 105could both be continuous and an additional layer could serve as a spacerin between.

Although routing of any complexity can be accomplished according to theart, it is not necessarily a complex matter to include routing for suchoptimal features as a running fluid for a temperature bath that iscompatible with high resolution microscopy. For example, referring toFIG. 27, which is a version of the FIG. 19 embodiment employingbilateral symmetry and a temperature bath, which in turn is a two-layerversion of the FIG. 6B embodiment, a channel pattern 107 is etched inthe upper 76 of two glass layers to provide for a running fluid 19 thatproceeds from an inlet 108 to an outlet 109 to establish a temperaturebath. Vias 110 may be etched in the upper layer 76 to aid in makingfluidic contact with fluid supply line connectors (not shown) atrespective inlet and outlet ends. To meet with standard requirements forhigh resolution microscopy, the bottom layer is preferably 170 micronsthick, with a refractive index of 1.515. The two glass layers aredirectly bonded without adhesive. The channel pattern 107 may be variedto accommodate other types of fluid supply line connectors; also,additional features (e.g., holes, notches, etc.) can be made in one ormore of the glass layers to aid in establishing such connections. Thechannel pattern 107 for the running fluid 19 can also be etched in thebottom glass layer instead, or both layers can be etched to enable agreater rate of flow for the running fluid 19 by providing a deeperchannel.

In general, optimal patterns for routing fluids in an FCA can bedetermined according to the art with the aid of fluorescent dyes (toobserve mixing and flow), optical doppler tomography (to detect flowvelocity), infrared microthermography (to determine temperaturedistribution), and other techniques (e.g., to measure volume, etc.); andso, the pattern examples described in this disclosure are intended forinterpretation as enabling examples, but not as limiting examples, aswill be appreciated by one skilled in the art.

Referring to FIG. 20C, it has been noted in the field of in vivomicroscopy that when wet immersion objectives are used (e.g., oil orwater), the microscope objective 84 will serve as a heat sink (orsource) unless it is at the same temperature as the physiologic mediumMM. This effect results in a loss of temperature control. WarnerInstruments, Inc. (Hamden, Conn.) notes that “it has been observed thatthe solution temperature directly above an immersion objective canchange as much as 10° C. for a 37° C. solution perfusing at 5 ml/min inan apparatus maintained at ambient [room] temperature [˜21-23° C.].”(Warner Instruments, Inc., “OW Series Objective Warmers,” Rev. Jan. 27,2004, p. 4). This effect would be especially disastrous in the case of aprenidial incubator because according to the traditional setup for highresolution microscopy using a wet immersion objective the objective 84would have to be placed directly underneath the patient resting in themicrocradle. In general, prior artisans in the field of in vivomicroscopy have responded to the heat sink problem either by warming themicroscope in a cage incubator or by heating the objective 84 by atemperature-controlled resistive means.

Okolab (Naples, Italy) employs a cage incubator for in vivo microscopyenclosing both the specimen and substantially enclosing the microscope.An exclusive reliance on this approach is inadequate because temperaturewill fluctuate when the cage is opened. Warner Instruments, Inc. employsa temperature-controlled resistive heating collar (the OW-37 ObjectiveWarmer) that heats an objective 84 indirectly by warming air very closeto the objective. Warner Instruments, Inc. claims that indirect warmingof this sort is less stressful for the objective 84 than a resistivemeans that warms by direct contact with the objective. A general problemwith employing a resistive means is that it may interfere with highlysensitive electrical or magnetic equipment used to investigatephysiological signals; also, temperature pull-down times are relativelyslow with this type of means because cooling relies on dissipation; butperhaps the most important and overlooked problem is that infraredradiation from a resistive heating element placed in close proximity toa prenidial infant may warm or overheat the patient in the incubator. Ofadded concern, a specific problem with heating air to warm the objective84 is that the heated air may rise to heat the cover glass CG by anundesirable amount.

When needed to eliminate a potential heat sink or source, a temperaturebath according to the invention is generally preferred to maintain adesired temperature of equipment such as a microscope objective placedin thermal contact with an FCA. To circulate a running fluid, theobjective may be surrounded by a water jacket, coils, or the like.Objective heater designs employing a copper tubing water jacket areknown in the art of live cell imaging. But preferably the temperaturebath may be integrally routed within a structure of the objective,particularly if the objective is fabricated using a technologycompatible with microfluidics, such as a layer-based technology.Microfabricated objectives of this sort are discussed further belowunder this sub-section on “Design Considerations”, including objectivesforming part of an FCA. Unlike a resistive means that cycles on and offto produce a desired temperature, the temperature bath does not subjectthe objective to a thermal cycle since the temperature bath is generallyconstant; in contrast, a resistive heating element is usually hotterthan a desired ambient temperature when turned on, and must therefore becycled on and off; thus, the temperature bath avoids subjecting theobjective to the stress of thermal cycling without relying on warmedair. If the temperature needs to be changed the temperature bath enablesfast pull-up/down times; the change can also be produced gradually;also, there is no electronic or magnetic signature, and there is noresistive heating element to generate infrared radiation in closeproximity to the infant. The temperature bath should circulate in such amanner (e.g., in a downward direction away from the patient) that agradient of heat transfer moves away from the patient care region.

In general, note that a degree of isolation between the patient careenvironment and the surroundings may still be desirable even despitebeing able to employ means to equilibrate temperatures. For example,popular manufacturers of high resolution microscopy equipment suggestoperating environments within a range of 20-30 degrees Celsius and ahumidity of 60% or less. This is outside the range of 37 degrees Celsiusand 95-100% humidity for prenidial incubation. Another problem isradiant heat transfer. In the art of neonatal incubation, double-walledincubators are employed to reduce radiant heat transfer between theinfant and the surroundings. For example, the Air-Shields® Isolette®C2000e infant incubator sold by Dräger Medical AG & Co. KG (Lubeck,Germany) employs a double-wall construction.

Referring to FIG. 28, the bottom of a microcradle may be formed by acomposite layer 111 comprising any number of layers. In this example, anintermediary layer 112 provides a thermal barrier or void 113 to isolatethe patient P from the surrounding environment 114 in the manner of adouble-walled incubator bottom. For example, the thermal barrier or void113 may comprise an air or vacuum chamber. An objective lens may bedesigned to accommodate specific parameters associated with the designof the composite layer 111. For example, the thermal barrier or void 113may be of a certain depth. Additional thermal barriers or voids may beformed by various techniques between or alongside channels or devices,around the sides of a vented microcradle, or elsewhere in an FCA. Suchtechniques may include the use of etching to create a void or the use ofstencil layers, layer spacers, or thermally insulating layers or regionsto produce thermal barriers or voids. Coatings, surface finishes, andmaterials with relevant properties may also be used to regulate anemission and absorption of thermal radiation. In general, FCA's may alsoinclude structures and devices to conduct, dissipate, or distribute heatfrom devices that warrant it (e.g., away from an integrated circuitchip).

A number of teachings in the field of microfluidics commonly assigned toNanostream, Inc. teach use of a stencil layer comprising channelboundaries cut in a pattern entirely through the layer to providelateral boundaries for fluid movement when sandwiched between top andbottom layers, e.g., the teachings of Pezzuto et al (U.S. Pat. No.6,418,968) and O'Connor et al (U.S. Pat. No. 6,481,453). The prior artof microfluidics does not teach or fairly suggest a substantially planarstencil layer to provide a thermally insulating layer in the form of athermal barrier or void when sandwiched between top and bottom layers;the prior art also does not teach or fairly suggest a thermal barrier orvoid etched in one or more layers to provide thermal insulation. Stencilpatterns are limited to open paths because separate cutouts will resultfor each closed path. However, various cutouts may be bonded to a layer,including in combination with a stencil, to achieve a desired pattern ina substantially planar layer including closed paths.

Microoptics is a rapidly evolving technology in microfabrication and itwould be highly desirable to have available an optical MEMSimplementation to replace bulky, large-scale microscopy and infraredmicrothermography for use with a vented microcradle. MEMS optics mayinclude such exemplary features as light sources, filters (e.g., acircular polarizer, interference filter, or tunable filter), condensers,compensators (e.g., a universal liquid crystal compensator or Pockelscell), mirrors, microlenses and microlens arrays, objectives, adaptiveoptics, diffractive optics, means of focusing, means of microscopicinstrument control (e.g., device positioning), means of signal detection(e.g., photodiodes or microbolometers), means of data communication,microinterferometry, microspectroscopy, microrefractometry, and more.But, generally speaking, a number of challenges still need to beovercome to develop optical MEMS implementations that on are on par withtheir large-scale counterparts. Also, integration of an optical MEMSdevice with a vented microcradle inside the fluid incubation mediumbrings with it additional challenges such as compatibility and devicecooling.

An optical MEMS implementation enabling a lateral optical path(side-to-side within the microcradle) for both microscopy (forvisualization) and infrared microthermography (for patient temperaturedetection) would be ideal. It would also relax the need for an opticallyclear bottom for the vented microcradle. However, in absence of such animplementation, a compromise route to optical MEMS would be to integrateMEMS optics with the clear bottom itself.

To give an example for illustration, Koninklijke Philips Electronics NV(Amsterdam, the Netherlands) makes an electrically focusable liquid lens(brand name FluidFocus) suitable for MEMS optics applications. Referringto FIG. 29A, a conducting aqueous solution 116 forms a convex meniscuswith respect to a non-conducting, immiscible, and hydrophobic liquid 117having a different index of refraction to form a lens; the aqueous 116takes on a convex shape in simple contact with a hydrophobic coating118. Referring to FIG. 29B, when a voltage difference is applied betweenrespective bottom and sidewall electrodes 119-120 a static charge buildsup, creating an attraction between cations in the aqueous andneighboring electrons residing on an outer surface of the sidewallelectrode 120 where it covers the inside of a glass wall 121; thisattraction compensates in a pseudo-hydrophilic sense for the otherwisehydrophobic coating 118; the attraction causes the aqueous to form aconcave lens shape. An insulator 122 separates electrodes 119-120.Bottom electrode 119 makes electrical contact with the conductingsolution 116 whereas sidewall electrode 120 is insulated. Notably, acapacitive relationship is formed between cations in the conductingsolution 116 and electrons residing on the surface of the side electrode120; it is a polar attraction of the aqueous for these cations thatcauses the sidewall surface to be wetted. The lens shape and, hence,focus varies with applied voltage; an applied voltage does not need tobe maintained to maintain the lens shape because the charges aremaintained by capacitance.

Kohashi et al (U.S. Pat. No. 4,030,813) teach a control element having aliquid layer attainable to a geometrically uneven state in response toan electrical signal, with function as an electrically controllableoptical lens. Berge et al (U.S. Pat. No. 6,369,954) teach a lens withvariable focus comprising a liquid lens (also known as a tunable liquidmicrolens) having a lens shape controlled by electrowetting. Berge et allicensee Varioptic, Inc. (Lyon, France) is the leading manufacturer offocusable liquid lenses. Modifications of the liquid lens concept areknown, e.g., the FluidFocus lens. For simplicity of comparison withrespect to various new modifications presented below, a discussion hasbeen given above of the Philips version (FluidFocus) because theFluidFocus lens employs straight sidewalls as opposed to slopedsidewalls employed by Varioptic lenses (e.g. ARTIC lenses), the latterbeing a little more complicated to manufacture. The Philips FluidFocuslens is disclosed in the published application of Feenstra et al (US2005/0113912). A comparable Varioptic lens is disclosed in the publishedapplication of Berge et al (US 2006/0126190).

New modifications of the liquid lens concept are needed to improvecompatibility of the lens with the present invention. For example,Phillips makes a FluidFocus lens having a diameter of 3 mm. However,lenses of substantially smaller diameter (e.g., 0.25-1 mm) will likelywork better with the present invention. But to achieve operability in asmaller diameter lens it becomes necessary to increase capacitancebetween the polar (hydrophilic) conducting solution 116 and the sidewallelectrode 120, otherwise the lens will not change shape or undesirablylarge voltages will be required to produce the change. Thus, one objectof modification is to promote increased capacitance. Various options cansatisfy this object to varying degrees. For example, the hydrophobiccoating 118 can double as an insulator 122 to strengthen capacitance byminimizing a distance between charges by eliminating an added thickness;moreover, a composition of matter to achieve this goal with as thin acoating as possible would be preferred. Another option is to employ aninsulator 122 with a high dielectric constant to promote capacitance.For the same reason, all else being equal, a hydrophobic coating 118with a higher dielectric constant will promote more capacitance than onewith a lower one. An effect on capacitance of tradeoff between acoating's thickness (i.e., charge separation distance) and the magnitudeof its dielectric constant is understood in view of capacitor physics.

In the drawing, note that although insulator coatings 122 andhydrophobic coatings 118 are being drawn and labeled separately toemphasize their distinct physical functions, it is understood that inpractice they may be one and the same coating. It will also beappreciated by one skilled in the art that a hydrophobic coating may beapplied wherever hydrophilic interaction is undesirable, and similarlythat a hydrophilic coating may be applied to promote hydrophilicinteraction; thus, an omission of such a coating in a part of thedrawing does not imply that such a coating would not be of use inspecific circumstances.

An approach to liquid lens design employing laminated planar layers ispreferred for use with the present invention because layer-based designsenable seamless integration with FCA designs. Philips and Variopticemploy laminated glass or plastic layers. For example, a FluidFocus lens3 mm in diameter and 2.2 mm in length employs an annular member for theglass sidewall 121 with glass end caps 123-124 to form a cylindricallens for use in a digital camera. (Philips Research Press Release,“Philips' fluid lenses bring things into focus,” Mar. 3, 2004.) However,the prior art does not employ microfluidics to fill the lenses. Instead,the prior art pairs together one drop each of oil and aqueous andsandwiches them between layers. A published application for a “DropCentering Device” by Berge (US 2005/0002113) underscores the complexityof this process. As taught by the present invention, a microfluidicapproach to filling the lens with liquid is needed for a very smalldiameter lens (e.g., 0.35 mm) and simplifies the filling of largerlenses as well.

FIGS. 30A-B show an exemplary small diameter liquid lens according tothe invention employing a layer-based approach compatible with an FCA;referring to FIG. 31A, a microfluidic channel 128 is etched in the topface of a middle glass layer 126 and a via is etched through the middlelayer 126 to form a microfluidic lens chamber 130; referring to FIG.31B, another microfluidic channel 129 is etched in the bottom face ofthe middle glass layer 126; though not shown in FIGS. 31A-B, and as bestseen in FIGS. 30A-B, the sidewalls of the microfluidic lens chamber 130are 1) coated with a conductor of electricity to form a sidewallelectrode (cylindrical capacitor plate) 120, 2) coated with anelectrical insulator 122, and 3) coated with a hydrophobic substance118; the insulator coating 122 and hydrophobic substance 118 maypreferably be one and the same coating; the coatings may be accomplishedusing conformal chemical vapor deposition; a conductive trace is routedto connect the cylindrical capacitor plate 120 to a point of electricalcontact; referring to FIG. 31C, a conductive trace 119 is deposited onthe top face of a bottom glass layer 127 to form a bottom electrode 119.

The three glass layers 125-127 are bonded together, e.g., with adhesive.As best seen in FIGS. 30A-B, the microfluidic channels 128-129 establishfluidic communication with the microfluidic lens chamber 130. To form aliquid lens, the microfluidic lens chamber 130 is first filled with anon-conducting, immiscible, and hydrophobic liquid 117 (e.g., an oil)via the microfluidic channels 128-129 and then a conducting, hydrophilicliquid 116 (e.g., an aqueous solution) is urged into the chamber 130 viathe bottom microfluidic channel 129; alternatively, the chamber 130 isfirst filled with the aqueous solution 116 and then the oil 117 isentered into the chamber 130 via the top channel 128. The water 116 andoil 117 touch each other so there is no gap in between and the meniscusbetween them forms a lens, the two liquids having different refractiveindexes. An exact filling of the lens chamber 130 to form a lens ispreferably accomplished using a visualization means. The visualizationmeans may include a microscope placed under the lens chamber 130 or aphotodiode array placed under the lens chamber 130. A test pattern orspecimen may be placed above the lens chamber 130 so that the progressof filling the lens chamber 130 can be monitored in terms of an imagevisualized and a degree of focus obtained. As shown in FIG. 30B, theliquid lens is shaped by changing a DC voltage applied between thesidewall electrode 120 and the bottom electrode 119. The sidewallelectrode 120 (insulated) is preferably given a negative charge and thebottom electrode 119 (in contact with the conducting solution) is givena positive charge, or vice versa. Focusing operations, e.g., shaping thelens by applying different voltages, or raising or lowering the meniscus131 between liquids (described below), may aid in monitoring the liquidlens filling process.

Referring to FIG. 32 in view of FIG. 30B, in addition to being able toform a very small diameter liquid lens, another distinct advantage ofemploying microfluidics to fill the lens chamber 130 of a liquid lens isthat a meniscus 131 between liquids may be lowered or raisedhydraulically to change a focal distance f between a lens and its focalpoint F by an incremental amount Δf. FIG. 32 shows oil 117 being urgedinto the lens chamber 130 via the top microfluidic channel 128 toincrease the focal distance by an amount Δf. Conversely, the aqueous 116can be urged into the lens chamber 130 via the bottom microfluidicchannel 129 to reduce the focal distance by an incremental amount. Inother words, the lens focus can be controlled not only by changing thelens shape by applying different voltages but also by moving the lenshydraulically within the lens chamber 130 by means of microfluidics.

Referring to FIG. 32, the middle layer 126 forming the lens chamber 130can be made of any thickness, thus enabling the meniscus 131 (liquidlens) to be moved up and down over any desirable range of distances.Because microfluidics can handle extremely small volumes of fluid and,hence, very small values for Δf, hydraulic focus of a liquid lensaccording to the invention enables an extremely fine focus, as will beappreciated by those skilled in the arts of hydraulics andmicrofluidics. A hydraulically focusable liquid lens may also becombined with another lens to provide zoom lens capability. In contrast,Varioptic relies exclusively on a reshaping of fixed liquid lenses toprovide a relatively limited ‘zoom’ focus.

Referring to FIGS. 33A-B, the aqueous solution 116 may be coated with athin layer of oil 117 to form a meniscus lens, with air 132 on the otherside of the meniscus. The shape of the meniscus lens may be changed byelectrowetting. (Note that a meniscus forms a lens even apart from anelectrowetting means.) Employing air 132 or other gas affords a degreeof thermal isolation within the lens chamber. Note that if both sides ofthe oil meniscus contain the same aqueous solution (as opposed toplacing air 132 on one side and aqueous 116 on the other) then the oil(non-conducting) layer 117 forming the meniscus will lie flat. Ofanalytical interest, deviations from flatness can be attributed todifferences in conducting solutions placed on either side of themeniscus formed by the non-conducting layer 117, such differences beingdetected as changes in focus caused by a curvature of the meniscus as afunction of voltage.

The simple meniscus lens shown in FIGS. 33A-B has a few limitations. Forexample, a pressure of the air 132 must be maintained to shape themeniscus by electrowetting, otherwise the aqueous 116 will move upwardbased on capillary action. But because the air 132 is highlycompressible, then even if the upper microfluidic channel 128 is valvedshut, there can still be some movement. Valving the lower microfluidicchannel 129 shut will eliminate net movement of the aqueous 116. Ingeneral, if channels such as microfluidic channels 128-129 are notclosed off or, more to the point, if a pressure opposing flow is notmaintained, then electrowetting forces will create a pumping effectbased on capillarity. In other words, though not shown in the drawing, ameans to resist undesirable flow is understood. Another limitation ofthis simple meniscus lens, as will be appreciated by one skilled in theart of optics, is that the respective shapes or radii of the convex andconcave sides of the lens cannot be independently controlled byelectrowetting.

FIGS. 34A-B show a more advanced liquid lens capable of assuming the sixbasic lens shapes: convex-concave, meniscus, biconcave, plano-concave,biconvex, and plano-convex. This versatile lens can easily bemanufactured in a simplistic embodiment employing five glass layers133-137. It may be noted in general that only layers in line with anoptical path need to be optically clear (e.g., top and bottom layers)and that the remaining layers can be of another material that is notnecessarily clear. Referring to FIGS. 34A and 35, microfluidic channels138-140 and vias 141-143 are etched in respective layers 134-136; thevias 141-143 combine to form an internal lens housing 144 (lenschamber). If desired to limit layer thickness, the channel 139 of themiddle layer 135 may be etched all the way through. Upper 134 and lower136 layers forming the internal lens housing 144 are identicallymanufactured except their top and bottom faces are oppositely oriented.The via 142 of the middle layer 135 is preferably etched wider than thecircumference of the internal lens housing 144 so as to provide a recess145 to help seat a lens formed by a hydrophilic, conducting liquid 116(e.g., an aqueous solution); the via 142 may also be crenulated toincrease surface area to promote hydrophilic interaction; a region 146of upper 134 and lower 136 layers is preferably hydrophilic to aid inseating the aqueous 116. Insulated sidewall electrodes covered by ahydrophobic coating are formed on the sidewalls of the upper 141 andlower 143 vias by depositing a conductor of electricity 120, insulator122, and hydrophobic coating 118. An uninsulated sidewall electrode 147is formed on the sidewall of the middle via 142 by depositing aconductor of electricity 120. Though not shown in FIG. 35, and as bestseen in FIG. 34A, electrical traces are routed from the sidewallelectrodes to a point of electrical contact. The upper and lowerportions of the internal lens housing 144 are filled withnon-conducting, immiscible, and hydrophobic liquids 117A-B (e.g., oils)and the middle portion is filled with a hydrophilic, conducting liquid116. In other words, the non-conducting liquids 117A-B do notnecessarily need to be the same. For example, as will be appreciated byone skilled in the art of optics, the non-conducting liquids 117A-B mayexhibit different refractive index dispersion. In operation, the shapesof upper and lower menisci 131A-B are independently controlled byapplying DC voltages between electrodes; as shown in FIGS. 34A-B, avoltage potential V1 applied to upper and middle electrodes controls theupper meniscus shape 131A and a voltage potential V2 applied to lowerand middle electrodes controls the lower meniscus shape 131B; V1 and V2observe the same polarity with respect to the middle electrode. As shownin FIG. 34A, fluid may be urged through top and bottom microfluidicchannels 138, 140 to provide fine changes in lens focus Δf. Fluid mayalso be urged via the middle microfluidic channel 139 to change thethickness of the aqueous layer 116 forming the lens.

Similarly, FIG. 36 shows a liquid lens paralleling the same design andoperation as the embodiment shown in FIGS. 34A-B, except the conductingand non-conducting liquid layers are reversed. Note that the routing ofelectrical contacts has been simplified to facilitate the drawing. Ineffect, the FIG. 36 embodiment joins two units of the embodiment shownin FIGS. 30A-B head-to-head and forms a single lens chamber 144. Likethe FIG. 34A-B embodiment, this lens is capable of assuming all of thebasic lens shapes. However, as illustrated in FIGS. 37A-B, anoperational difference needs to be pointed out. In FIG. 37A, conductingliquids 116A-B are exposed to a DC voltage of the same polarity using apotential difference that for illustrative purposes causes respectivemenisci to lie flat. But then, referring to FIG. 37B, when the polarityis reversed on one of the liquids, in this case conducting liquid 116B,a “meniscus capacitor” is formed by a build up of charges of oppositepolarity on either side of the non-conducting liquid 117, therebyresulting in a biconcave lens shape. It is contemplated that this sortof meniscus capacitor will enable a number of analytical advancements,including a study of an effect of ion concentration and ion species onrefractive index and refractive index dispersion; additionally,properties of the conducting 116A-B and non-conducting 117 solutions canalso be investigated analytically by detecting changes in focus causedby changes in the curvature of the menisci as a function of voltage.

Referring to FIGS. 38A-B, any number of liquid lenses LI, L2, . . . , LNmay be combined to form an objective lens. The objective may be separatefrom a cover glass or, as shown, the cover glass CG may form the toplayer of the objective. In some cases, as shown in FIG. 38B, if thefront lens is not formed by an aqueous solution then it may be desirableto place a liquid layer with the same refractive index as the aqueoussolution between the front lens L1 and the cover glass CG to account forrefractive index requirements.

In the prior art, a limitation arises because in some cases it isdesirable to focus at some depth into an aqueous medium but in othercases it is desirable to focus on cells plated on the surface of thecover glass CG. According to the art of high resolution microscopy, thiswould entail having to switch between water immersion and oil immersionobjectives. But switching immersion fluids would be particularlycomplicated and problematic with in vivo microscopy because theswitching process would involve contact, e.g., to remove water or oil,which in turn would have thermal implications for the in vivoenvironment. The present invention is able to solve this problem with amicrofluidic approach. In general, in some cases it may be desirable toexchange a liquid in a liquid lens objective via microfluidics, e.g., toemploy a liquid with a different index of refraction; for example, toimage coculture cells plated on the surface of the floor of a ventedmicrocradle (the floor of the microcradle forming the cover glass CG ofthe objective), it may be desirable to employ a liquid immediately underthe cover glass CG having the same index of refraction as the coverglass CG, e.g., an oil with a refractive index of 1.515; in contrast, tofocus at some distance past the cover glass CG into the aqueous solutionor patient's body, it may be desirable to employ a liquid immediatelyunder the cover glass CG having an index of refraction similar to theaqueous solution or the tissues of the body, e.g., a refractive indexbetween 1.33 and 1.39.

Referring to FIGS. 20C and 39, according to theory in microscopy,numerical aperture, which is a measure of an objective's light gatheringability and resolution power, is limited by the formula n·sin(θ) where nis the refractive index of an immersion fluid between the cover glassand the objective front lens and θ is one-half the angular aperture. Thevalue of θ and, hence, numerical aperture increases as the workingdistance WD decreases. Referring to FIGS. 38A-B, a liquid lens objectiveaccording to the invention enables an extremely small working distanceby incorporating the cover glass CG with the objective itself; all elsebeing equal, this serves to benefit an increase in numerical aperture.

But it should also be noted that the formula for numerical apertureassumes a flat cover glass, as shown in FIG. 40. However, referring toFIG. 41, a microlens cover glass can be used to improve numericalaperture and/or reduce the dependency of numerical aperture on therefractive index of the immersion fluid between the objective front lensand the cover glass. A microlens cover glass is not practical with priorart objectives because it would be very difficult to align the microlensalong the central axis of the objective if the objective lens and coverglass are separate, as is the case with traditional microscopy. Thisproblem is overcome by integrating the cover glass CG with theobjective, as shown in FIGS. 38A-B. Notably, however, an alignmentmeans, e.g., using automation combined with machine-readable indicia ofalignment, would also make it possible to use a microlens cover glasseven without such integration.

Lenses of a layer-based objective according to the invention do not needto be exclusively liquid lenses, for an objective may also beconstructed in layers using any combination of microfabricatedmicrolenses as well. For example, referring to FIG. 41, a layer-basedobjective may include a microlens cover glass 148 and a microlens frontlens 149, and the space between the front lens 149 and cover glass 148may be filled with air 150 to provide a thermal barrier. When focusingon a point on the surface of the cover glass, use of the plano-convexmicrolens cover glass 148 compensates for a loss in numerical aperturethat would otherwise occur with an oil immersion objective whensubstituting air as the immersion medium.

Those skilled in the art of microscopy will appreciate that a microlenscover glass may employ any variety of lens shapes depending on suchfactors as the design of the objective (and particularly the frontlens), the refractive indices and refractive index dispersions involved,and whether focus is aimed at the surface of the cover glass or past thecover glass into tissues or through a medium; for example, a biconcaveor positive meniscus microlens cover glass may be followed by a biconvexobjective front lens.

Referring to FIG. 42, a patient P is resting on the floor layer of aside-vented microcradle with a clear “cover glass” bottom CG. The coverglass CG may be a microlens cover glass. A microlens may be directlyformed in the floor layer or microlens layers may be laminated to thetop and/or bottom of the floor layer. Additional FCA layers 151 maycontain any combination of exemplary features of an MEMS optical system,including but not limited to thermal barriers, temperature baths,microlenses, liquid lenses, traditional lenses, prisms, beamsplitters,polarizers, filters, mirrors, and image receptors (e.g., a CCD, CMOS, ormicrobolometer).

Additionally, or alternatively, optical equipment 152 separate from theFCA may also be placed underneath the FCA of a vented microcradle.Compared to traditional optical systems, a particular advantage of alayer-based system employing micro fabrication technology is that it canbe readily practiced in array form. Referring to FIGS. 5 and 43, alayer-based optical system may be practiced in array form in the contextof FCA layers 151, so that optical devices (e.g., objective lensesOL1-OL3) may be focused not only on the vented microcradle but also onvarious other FCA devices as well. Referring to FIG. 44, optical devices152 placed underneath (or above) the FCA may also be practiced in arrayform so that individual devices (e.g., objective lenses OL1-OL3) can beselectably moved into position mechanically.

From an optical perspective, referring to FIG. 30A for example, the lensformed in this case may be thought of as a lens doublet, given that thetop 117 and bottom 116 lenses may have different refractive indexdispersion properties. Similarly, referring to FIG. 34A for example, thelens formed in this case may be thought of as a lens triplet, given thatthe top 117A, middle 116, and bottom 117B lenses may have differentrefractive index dispersion properties. Referring to FIGS. 38A-B,liquids with various optical properties, such as refractive indexdispersion, may be employed according to the art, for example, tocorrect the objective for chromatic aberration. Cargille-SacherLaboratories, Inc. (Cedar Grove, N.J.) specializes in optical liquids(and gels) of desirable properties. Other substances, e.g., proteins,may also find interest.

To make a highly corrected immersion objective, such as a planapochromat objective, opticians must take into account the refractiveindex and Abbe's number of each lens element comprising the objective.The Abbe's number is a measure of refractive index dispersion. Referringto FIG. 45, in order to allow for more flexibility in choosing liquidswith specific optical properties, a region in a liquid objective thatwould otherwise be occupied by a single liquid may be partitioned with atransparent layer 153 so that two liquids with different opticalproperties (LIQ. A and LIQ. B) can occupy opposite sides of thepartition.

Referring to FIG. 46, and to compare with FIG. 37B, ionic charges may beplaced on both sides of a microlens 154 using a voltage potential Vapplied to liquids on opposite sides of the microlens 154. At thesurface of the microlens, this can affect optical properties related toion species type and concentration, such as refractive index andreflectance. Ion concentration and ion species type can be varied inLIQ. A and LIQ. B as desired, as can voltage potential V. However,charge amounts on respective surfaces of the microlens will be equal andopposite, and capacitance between the charges (and, hence, local ionconcentrations) will depend on the thickness of the microlens, which isnot uniform, and the dielectric constant of the microlens material,e.g., glass. Referring to FIGS. 47A-B, a more versatile arrangement isobtained by separately coating each side of the microlens 154 with aconductor of electricity 155 covered by an electrical insulator 156 toform separate capacitors with respect to ions in the neighboringliquids. Like the electrical insulator 156, the conductor of electricity155 (e.g., indium tin oxide) must also be transparent. By applyingvoltages V1 and V2, ionic charges on either side of the microlens can beindependently controlled. In general it may also be noted that amicrolens may be coated with any variety of optical coatings.

FIGS. 48A-E depict human anatomy at different stages of prenidial life,that is to say, during life before implantation. Referring to FIG. 48A,a baby is conceived within an egg; three small polar bodies are notshown. At this point the baby's body incorporates a cell having a cellsize much larger than usual. This is the early embryo stage. The cellsize will be reduced by successive divisions. The first few celldivisions proceed linearly in a serial fashion (1, 2, 3, . . . , 8)rather than exponentially in a parallel fashion (1, 2, 4, 8) as was oncemistakenly presumed. A mulberry-shaped cluster of cells (the “morula”)will develop. Before leaving the protection of the shell-covered eggcapsule, the baby must first grow a protective spacesuit. Referring toFIG. 48B, by the late embryo stage, the baby is surrounded by aspacesuit filled with fluid. The baby is attached to the inside wall ofthe spacesuit. From the time when the baby first starts to build aspacesuit until after birth when the umbilical cord is cut, the baby'swhole body consists of a “baby” part and a spacesuit part. The spacesuitportion of the anatomy is called the peripheral body and the astronautor baby portion is called the formal body. The entire body is called theconceptus, which is another name for the baby during gestation. In laterdevelopment the spacesuit includes an umbilical cord and a plug to plugthe spacesuit into the maternal body. At hatching time, the spacesuitonly consists of a covering. The covering takes the form of a sphericallayer of relatively large cells and is filled with fluid, in contrast tothe smaller cells of the baby's formal body. No cord or plug is formeduntil after implantation begins.

Referring to FIG. 48C, a baby uses specialized shell-breaker cells onthe covering of his or her spacesuit to make a hole in the shell of theegg, and then he or she exits the egg through the hole in an extrusivebehavior involving movement. The baby relies on brain power in the formof molecular computing inside cells to implement this intelligentbehavior, and the basis for locomotion is contained in the form ofchemical contractions exerted on the cytoskeleton within cells, as isall muscular movement. Human babies hatch from their eggs about 5-6 daysafter fertilization. Referring to FIGS. 48D-E, after hatching the babyleaves the empty egg capsule behind and is now free to approach thelining of the maternal uterus and implant. The baby's formal body isprotected from exposure outside the egg capsule thanks to the protectivespacesuit the baby put on before leaving the egg capsule.

The significance of implantation is simply that this is when the babyattaches his or her spacesuit to the mother ship, so to speak, like anastronaut would. Referring to FIG. 48E, note in particular that thetissues of the baby's spacesuit are bare because there is no more shellsurrounding the baby's body. The reason for the baby's protectivespacesuit is that without it the tissues of the formal body (the “baby”part) would be exposed inside the mother.

According to the process of implantation, a plug formed by chorionicvilli on the outside of the spacesuit (at the far end of the umbilicalcord) attaches the baby's spacesuit (and, hence, the baby) to themother's body for the remainder of gestation. The baby exits thespacesuit after the covering (the birth sac) breaks. The spacesuit willbe discarded after birth when the umbilical cord is cut. Then theastronaut goes home in a car seat from the hospital.

It took doctors a long time to realize this. They used to think thebirth sac and umbilical cord were a part of the mother's body, not thebaby's. Another problem preventing their grasp of the significance ofimplantation was that they thought an egg simply attached to thematernal body. They did not realize the baby has to hatch first!Accordingly, they did not realize that the field they called comparativeplacentation is really the subject of gestational spacesuits and theirmanner of attachment. Notably, different species exhibit different typesof gestational spacesuits, which also form different types ofattachments to the mother.

One skilled in the art of MEMS will appreciate that thermistors may beplaced in contact with an embryo by means of a MEMS-based device orprobe to monitor patient temperature. An array of thermistors may beplaced around the embryo to monitor thermal gradients. A contact meansof temperature monitoring other than a thermistor, e.g., a thermocouple,may also be employed. Unlike embryos, who are covered by the shells oftheir eggs, the body tissues of hatchlings are directly exposed. Forthis reason, a contact means presents an added complication whenmonitoring the temperatures of hatchlings.

Attention is therefore turned to the subject of non-contact temperaturemeasurement.

Referring to FIG. 49, the spectrum of electromagnetic radiation isdivided into gamma ray, X-ray, UV (ultraviolet), visible, infrared,microwave, and radio wave spectral regions, in order of increasingwavelength, decreasing frequency, and decreasing photonic energy.Referring to FIGS. 49-50A, a visible spectrum defines a region of theelectromagnetic spectrum to which the human eye is sensitive. Thewavelength of light in the visible region ranges from 400 to 700nanometers for most eyes, but some people's eyes are sensitive towavelengths as low as 380 nm or as high as 780 nm. FIG. 50A divides thevisible region by color response for the human eye. Referring to FIGS.49 & 50B, infrared light extends from the longest wavelength of visiblelight (red light), at approximately 700 to 750 nanometers, all the wayto the shortest microwaves at 1000 microns. FIG. 50B divides theinfrared region using an accepted scheme, though a number of otherschemes are also known and may rely on similar terms.

Radiometry is the general science of detecting and measuringelectromagnetic radiation as it is emitted and reflected by physicalbodies. The term radiometer refers broadly to any means of detecting andmeasuring electromagnetic radiation. Bodies naturally emit a broadspectrum of electromagnetic radiation as a function of theirtemperature, with a shift towards radiation at higher frequencies(shorter wavelengths) with increasing temperature. In simple terms, thetemperature dependence of this phenomenon enables the temperature of abody to be measured in a non-contact way by means of a radiometer.

In this disclosure, emissive flux means an energy amount of radiationemitted over a given surface area per unit time (dimensions: Watts permeter squared). Elsewhere, emissive flux may be known as radiantemittance or radiant flux per unit area.

A number of design considerations need to be factored in when using aradiometer for the non-contact temperature monitoring of a human embryoor hatchling. First of all, the emissive flux of the patient's body willbe greater at certain wavelengths than at others. Second, media betweenthe patient's body and the radiometer will generally absorb radiationmore strongly at certain wavelengths compared to others. Third of all,the sensitivity of the radiometer to radiation may vary according towavelength depending on the radiometer's physics.

Referring to FIG. 51, a Planck's law distribution of perfect black bodyradiation describes radiation amount as a function of wavelength for aperfect black body at 37 degrees Celsius. According to Wien'sdisplacement law, a maximum radiation amount is emitted at a wavelengthof 9.3 microns for a perfect black body radiating at 37 degrees Celsius.

Without considering absorption by intervening media, such as by thepatient's incubation medium, and without considering the physics of theradiometer, a radiometer that is sensitive to wavelengths near 9.3microns, assuming this is where emissive flux is highest at 37 degreesCelsius, would seem to provide the best alternative. For example, a QWIPinfrared camera is sensitive to radiation in the 8-9.2 micron range. Tocompare, an indium antimonide (InSb) infrared camera is sensitive in the3-5.5 micron range, an area of the perfect black body distribution curveat 37 degrees Celsius where emissive flux is significantly lower atgiven wavelengths. “This [comparison] can be misleading,” however,“because QWIPs have much lower quantum efficiency (ability to turnphotons into charge) than InSb detectors,” says Ross Overstreet, asenior scientific engineer at FLIR Systems, Inc (Wilsonview, Oreg.).This example helps illustrate why the physics of the radiometer (e.g.,quantum efficiency, noise) must be considered.

Absorption by intervening media must be considered as well. FIG. 52A isa graph of the absorption coefficients for liquid water as a function ofwavelength. Although water is highly transparent in the visible regionof the spectrum, water absorption increases significantly beginning at1450 nm in the infrared region. FIG. 52B focuses on absorptioncoefficients for liquid water in the 1-10 micron range. Although stillrelatively high, water absorption is significantly less in the 3.5 to5.5 micron range (where InSb detectors respond) as compared to the 8-9.2micron range (where QWIP detectors respond). This implies that althougha perfect black body radiating at 37 degrees Celsius emits more stronglyin the 8-9.2 micron range than in the 3.5-5.5 micron range, afterabsorption by an intervening aqueous medium a stronger emission may beactually be perceived by a detector operating in the 3.5-5.5 micronrange.

Referring back to an exemplary FIG. 24A, absorption is also an importantconsideration when a layer 95 forming an optical window 90 has toprovide for transmission over diverse wavelengths. For example, quartzglass (SiO₂), though clear in the visible region, only provides opticaltransmission to about 3.7 microns in the infrared. Thus, an opticalwindow made of quartz glass will not provide a clear path for a QWIPcamera, because quartz glass is opaque in the whole range (8-9.2microns) of a QWIP's sensitivity.

Optical windows with various transmission ranges are known in the art.When employing a material as an optical window for a microcradle, anumber of considerations must be taken into account in addition to thetransmission range itself. These include but are not limited to anability to make the optical window thin (e.g., as a cover glass 170microns thick), an ability to form lenses, an ability to accept apolish, solubility, scratch resistance, a need for anti-reflectioncoatings due to high index of refraction, biocompatibility, an abilityto accept surface coatings to enable biocompatibility, modulus ofrupture, and an ability to be modified to extend transmission range.Such considerations will be appreciated by those skilled in the art.

The ability of electromagnetic radiation to pass through a material atgiven wavelengths is known generally as transmission. Transmission isquantified by those skilled in the arts of optics and spectroscopy usinga number of related measurement techniques, such that the material canbe variously defined as having a low absorbance, low transmission loss,high transmittance, low absorption coefficient, low extinctioncoefficient, high penetration depth, or low optical density. Externaltransmission takes into account reflection at the surface of thematerial as well as internal transmission. For materials with a highindex of refraction, reflection losses can be high unlessanti-reflection coatings are used. Internal transmission takes intoaccount both absorption and scattering. Note that for emissions by abody submerged in a fluid only the internal transmission data for thefluid medium are pertinent, not the external transmission data for thefluid.

Referring to FIG. 52B, in the mid wavelength IR (MWIR) region (3-8microns) water absorbs the least at 3.80 microns. Those skilled in theart of optics are able to engineer coatings to reduce reflection lossesfor a selected wavelength or wavelengths. For example, as infraredradiation emitted by a patient's body travels through an aqueous medium,it is desirable that wavelengths to which a radiometer is sensitive arenot reflected back at the surface of an intervening material such as thecover glass bottom of a microcradle. By taking into account the emissiveflux of the patient at given wavelengths, subtracting internaltransmission losses in the aqueous medium, and subtracting internaltransmission losses in the cover glass material, one determines thewavelengths that would potentially yield the most intense signal ifthere were no reflection losses associated with the cover glassmaterial. Thus, in view of the wavelengths to which the radiometer issensitive, those skilled in the art of anti-reflection coatings will beable to coat the cover glass material to minimize such reflectionlosses.

In a white paper titled, “Common Misconceptions Related to InfraredInspection Ports,” Martin Robinson, managing director at GlobalMaintenance Technologies (Chelmsford, United Kingdom), explains thathigher transmission in an optical window is not necessarily of greateradvantage to thermography if it comes at the expense of other desirableproperties. This is because a radiometer can be calibrated to accountfor, say, a 75% transmission loss as easily as it can for a 50%transmission loss. This point is particularly important since somesuppliers of optical materials specify a transmission range that isnarrower than a range including relatively weak transmissions astransmittance falls off. In other words, to detect and measuretemperature a radiometer only needs enough incident radiation todistinguish temperatures.

For example, referring to FIG. 4, placing a thermal imaging camera 21above the patient P has the disadvantage that thermal radiation from thepatient will be substantially absorbed by an intervening layer ofaqueous medium M above the patient P. However, the camera 21 onlyrequires enough radiation from the patient P to distinguish her or histemperature from the temperature of the surrounding medium M. Theability to make this distinction will be aided if the microcradle isopaque and reflection back to the camera is limited. In other words, inpractice the camera only has to be able to distinguish between variousradiation amounts.

To give another example, when used in thin layers many materials willtransmit beyond what may typically be quoted as their useful range ofspectral transmission. For example, a 170 micron layer of IR grade fusedsilica (SiO₂) will allow for appreciable transmission at 3.80 microns, aspectral region where MWIR water absorption is at a minimum. In otherwords, although the MWIR transmission of fused silica, for example, isinferior to that of a number of other optical window materials, as longas the material permits sufficient transmission so that a givenradiometer can perform useful thermography, then the suitability of thematerial for other concerns (e.g., visible light transmission,biocompatibility, etc.) will dominate.

Those skilled in the arts of optics and spectroscopy will appreciatethat there are a large number of optical window materials availabletoday, with new materials under development. Sapphire (Al₂O₃), calciumfluoride (CaF₂), zinc sulfide (ZnS), magnesium fluoride (MgF2), and zincselenide (ZnSe) name but a few. Such materials can generally be providedwith coatings to achieve improved biocompatibility, durability, or tominimize reflection losses. FIG. 53 shows an external transmissionreport for uncoated sapphire, which is relatively transparent in boththe visible region and in the 3-5.5 micron infrared region of InSbsensitivity.

With respect to the clear bottom 3 of a side-vented microcradle thatserves as an optical window, the optical window can be pieced in (e.g.,layer 95 of FIG. 24A), used continuously (e.g., layer 34 of FIG. 6B), ormay include microfabrication (e.g., the clear bottom layer 3 of FIG. 23,which contains etched microchannels for a running fluid 19). PrecisionMicroFab, LLC is skilled in the art of laser milling microchannels insapphire.

Referring to FIGS. 54A-B, optical access to a patient P in spectralregions of strong water absorption (e.g., in infrared regions past thenear infrared) is preferably accomplished via the clear bottom 3 of aside-vented vented microcradle. In this case, the clear bottom 3 isformed of a layer material 157 that transmits electromagnetic radiationat desirable wavelengths. Referring to FIG. 54A, during the embryonicstage of life the patient P is bounded by the inside boundary 158 of theshell 159 of the egg capsule 160. In this example, the baby's egg 160 is120 microns in diameter and the shell 159 is 10 microns thick. Thus, theinner dimension comprising the tissues of the body of the patient P is100 microns in diameter. As shown in FIG. 54A, due to the sphericalgeometry of the egg 160, nearly 70% or more of the patient's body insidethe egg 160 will be observable via electromagnetic radiation that passesvertically through a thin layer 161 of aqueous incubation medium M nomore than 10 microns thick.

Referring to FIG. 54B, an absorption of infrared radiation by theaqueous medium M can be significantly reduced by means of a concavemicrolens 162 that has been formed in the layer material 157 forming theclear bottom 3 of the vented microcradle. As shown in FIG. 54B, theconcavity of the microlens 162 substantially fits the spherical shape ofthe egg 160 to the effect that there is little medium M left between theegg 160 and the layer 157 that serves as an optical window. In this way,absorption of electromagnetic radiation by an intervening aqueous layeris substantially avoided. Noted is that although the bottom side 163 ofthe layer 157 forming the optical window is shown to be flat in FIG.54B, thus forming a plano-convex microlens, a biconcave orconvex-concave (meniscus) microlens shape may also be practicedaccording to the art of optics. As shown in FIG. 54B, the concavity ofthe microlens 162 is slight enough that exposure of the patient P toventilating fluid medium M is not substantially occluded; moreover,stagnation of a portion of the patient's body is not an issue in thisrespect inasmuch as the patient P exhibits a gradual turning inside theegg 160 (somersaulting behavior).

In this disclosure, emissivity (also known as emittance) is a measure ofa body's emissive flux (or radiation amount) at a given temperaturerelative to the emissive flux (or radiation amount) of a perfect blackbody. Emissivity is a dimensionless ratio ranging from 0 (no emissiveflux) to 1 (emissive flux of a perfect black body). In other words, abody with an emissivity of 0.75 emits 75% as much radiation as a perfectblack body. Human skin has an emissivity of 0.98, which approaches veryclosely that of a perfect black body.

Unlike a perfect black body, ordinary bodies and substances do notexhibit a smoothly continuous distribution of radiation amount as afunction of wavelength; instead they exhibit a “fingerprint” patternthat characterizes the substance. Such a pattern occurs because asubstance tends to emit radiation at the same wavelengths it absorbsradiation. For this reason, the emissions spectrum of a substance tendsto mimic the absorption spectrum. Looked at from the perspective oftransmission, the substance will tend not to emit so strongly atwavelengths where it transmits. In contrast, according to definition aperfect black body absorbs and (hence) emits radiation at allwavelengths. Consequently, when bodies or substances are compared to aperfect black body it is understood that emissivity regards either anaverage comparison over a range of wavelengths or a comparison at agiven wavelength.

“The emissive characteristics of a target can be quantified byevaluating its relative emittance at two known wavelengths or at twoknown temperatures,” explains Pat Finney, Senior Applications Engineerat FLIR Systems, Inc. (Jacob, “Thermographic Imaging Essential to AvoidThermal Calamities,” Evaluation Engineering, August 1996.) To estimateemissivity for human embryos and hatchlings the two wavelength approachcan be applied for a live determination and the two temperature approachcan be applied for an autopsy determination. Similarly, the twowavelength approach can be applied to estimate emissivity forunfertilized eggs, but unfertilized eggs subjected to temperaturechanges may be made unhealthy.

In the context of an incubator system for human embryos and hatchlings,U.S. Pat. No. 6,694,175 teaches the need to distinctly “measure anactual body temperature of the human embryo or hatchling” for the reasonthat “said body temperature can differ from an ambient temperature of afluid incubation medium”. (Claim 1) Prior to this teaching practitionersinvariably confused ambient temperature readings as if they representedaccurate indications of patient temperature. This error has beenpersistent even despite the patented teaching.

In the general case of non-contact temperature detection and measurementby means of a radiometer, there are many unknown variables toaccommodate. For this reason, in general even a well calibratedradiometer will have difficulty in determining an actual temperature toan accuracy of 0.1 Celsius degrees. This is true even of a relativelyspecialized radiometer such as an ear thermometer, because variablessuch as ear shape, placement of the thermometer, and wax in the ear canaffect readings.

From an engineering perspective, however, use of radiometry to measurethe temperature of a human embryo or hatchling in a non-contact waypresents an idealized scenario: Parameters of the care environment(e.g., temperature, composition of the fluid incubation medium, etc.)are highly controlled, the patient's body is largely spherical and isthus amenable to simplified calculations, the distance between thepatient and the radiometer can be known with micron accuracy, and theemissivities of the patient and various constituents of the careenvironment can all be determined along with the absorbances ofintervening media.

Further idealization is provided in that the care environment canaccommodate references for calibration purposes, including thermistorsand emissivity test patterns. In the usual case of the prior art ofthermography, a radiometer is calibrated separately from a taking oftemperature measurements, and then temperature measurements are takenafterwards in the field. However, in addition to performing this type ofcalibration procedure, a care environment according to the invention maymaintain calibration references in the radiometer's field of view.

For example, a thermistor may be placed in the radiometer's field ofview on the floor of the microcradle. The thermistor itself may becalibrated in advance by submerging the floor layer in a temperaturebath prior to assembly. Hart Scientific, Inc. (American Fork, Utah)provides high accuracy temperature baths suitable for this purpose.Furthermore, a thermistor surface facing the radiometer may beconditioned (e.g., by coating) so that it exhibits a known emissivity(e.g., 0.98). In this way the temperature and emissivity of an areacovered by the thermistor will be accurately known so as to serve as areference. Note that if the temperature of the fluid incubation mediumis accurately known, an area of known emissivity at the temperature ofthe fluid medium will suffice without the thermistor. Either way, whenthe radiometer focuses on the reference area, the radiometer's responseto emissions from that area will be calibrated based on the presence ofa known reference for temperature and emissivity.

Emissivity test patterns placed in the radiometer's field of view canalso be employed. Referring to FIG. 55, an emissivity pattern containinga plurality of areas having known emissivities and transparencies may bedeposited on a surface in the radiometer's field of view. Opticalfilters may also be used so that the patterns are restricted to certainwavelengths.

Referring to FIG. 56, a more specific example of an emissivity patterntakes advantage of selective emission by different substances C1 and C2.Consider a radiometer that is sensitive to radiation in the 3-5.5 micronrange. Referring to FIGS. 57A-B, a coating substance C1 is relativelytransparent in the 3-5.5 micron range, except at 3.5 microns where itstrongly absorbs; similarly, a coating substance C2 is also relativetransparent in the 3-5.5 micron range, except at 5.3 microns where itstrongly absorbs. Referring to FIGS. 58A-B, because C1 and C2 absorbradiation at respective wavelengths they will also emit radiation atthese wavelengths; the radiation amount associated with these emissionswill vary according to temperature. Referring back to FIG. 51, becausethe slope of the distribution curve is changing at different rates fordifferent wavelengths, the ratio of radiation amounts for C1 and C2 willchange according to temperature. Importantly, a plot of this ratio as afunction of temperature will serve as a calibration reference. In otherwords, by comparing radiometer readings for areas C1 and C2 and takingtheir ratio, a corresponding temperature can be looked up by referringto a plot of this ratio as a function of temperature. Greater resolutionwill be obtained by choosing C1 and C2 at given wavelengths such thatthe rate change in radiation amount per unit temperature is changingdisparately between them, as can be seen from an examination of thedistribution curve, because in this way the change in ratio per unittemperature will be greater than if the rate change in radiation amountwere similar between the two. A clear area may be maintained as areference for subtracting background emissions; filters for variouswavelengths can also be employed to enhance resolution. Interpolationmay also be employed. In theory, C1 and C2 could also be chosen suchthat they transmit in a specific region and absorb elsewhere, which isthe opposite of the situation depicted in FIGS. 57A-B.

In the prior art of thermography, to access, for example, a high voltageelectrical panel with a thermal imaging camera, an IR window is placedin the door that covers the panel so that the panel can be viewedwithout exposing the operator to high voltages. Then, in order todetermine temperatures, the transmittance of the IR window must beentered as a parameter. In contrast, by employing an emissivity patternaccording to the FIG. 56 embodiment of the invention, there is no needto know the transmittance of the IR window because the ratio of C1 andC2 will not be affected. Thus, whether affixed or patterned on an areaof the floor of a microcradle, another area of an FCA, or on an area ofan electrical panel, an emissivity pattern according to the FIG. 56embodiment offers a uniquely flexible means of calibrating a radiometersuch as a thermal imaging system for real temperature. For example,instead of having to worry about transmittance changes when interveningoptics or materials are changed, the ratio of C1 to C2 can be reliedupon for temperature calibration. (An exception occurs when anintervening material selectively transmits at the wavelengths of C1 andC2.)

In general, an emissivity pattern may have a clear backing, a reflectivebacking, or a backing of known emittance or emittances, in anycombination.

Calibration test patterns may be endowed with machine-readable indiciaso they can be recognized and interpreted by a computer.

In general, software may be used to aid in temperature calibration,determination, and interpretation. FLIR Systems, Inc. is a provider ofsoftware products to acquire, calibrate, process, analyze, and storedata from digital infrared camera systems.

Because field-of-view calibration of a radiometer is possible, U.S. Pat.No. 6,694,175 teaches use of an infrared camera attached to a cameraport of an inverted microscope to thermally image a patient. In such acase, however, it should be noted that the microscope's optics mustsupport infrared wavelengths to which the camera is sensitive. Fluoriteoptics have the advantage of being fairly standard in the microscopyindustry while at the same time being able to transmit both visible andinfrared light. Accordingly, an inverted microscope equipped with dualcamera ports can be used to thermally and visually image the patient,such that one camera port is used for a CCD or CMOS camera to capturehigh resolution visible images while the other is used to support anInSb, QWIP, or other infrared camera for thermal imaging.

Thermopile sensors are contemplated for use in non-contact temperaturemeasurement in a prenidial care environment. Widely used for earthermometers, thermopiles consist of a series of thermocouples, suchthat a reference side is exposed to a reference temperature and anobject side is exposed to incident radiation emitted by a target. Sincea precise temperature bath is used in regulating the care environment,the reference side can be controlled by the same bath. Unlike InSb andQWIP infrared cameras, thermopiles can detect a broad spectrum ofradiation, although filters can be employed to restrict sensitivity to adesired range. Thermopile technology is highly cost competitive, runningin the hundred dollar range as compared to InSb and QWIP technologyrunning in the hundred thousand dollar plus range.

It may be noted that a sensitive enough radiometer will be able todetect temperature even with respect to wavelengths that regard verylittle emission at 37 degrees Celsius. A radiometer sensitive enough todetect temperature using wavelengths below 1450 microns, where strongwater absorption begins, would offer extreme versatility in terms of anability to position the radiometer with respect to the patient. Incontrast, a radiometer functioning at wavelengths of strong waterabsorption will prefer optical access to the patient either from thebottom surface or from another surface in very close proximity to thepatient.

In addition to providing optical access to a patient through the clearbottom 3 of a vented microcradle in a substantially vertical direction,optical access to the patient may also be gained through the flooring atan angle, including with the aid of mirrors, lenses, prisms, fiberoptics, specialized optical windows, and so on. Optical access may besimilarly gained from the sides or above, or through an alternatelyrouted location in the flooring.

Inasmuch the clear bottom 3 of a side-vented microcradle provides apreferred route for optical access to a patient, competition by visibleand infrared equipment will be somewhat constrained. Accordingly, asharing of the optical route by means of prisms, beamsplitters, dichroicmirrors, dual camera ports, and so on may be necessary. To improve orshare access, it may also be necessary to move equipment, or the FCAitself, by means of a motorized x-y stage or slider, for example.

Based on strong water absorption, a liquid lens employing water as alens liquid would not be optimum for transmission in an infrared regionabove 1450 microns. However, those skilled in the arts of optics andspectroscopy will be able to select suitable optical liquids accordingto the following simple method. In general, by means of spectroscopythose skilled in the art can determine the transmission losscharacteristics of a material at given wavelengths. Other opticalparameters such as the index of refraction and Abbe's number can also bedetermined. Referring to FIG. 59, having found liquids with suitableoptical properties, optical liquids 1 & 2 will be compatible for forminga liquid lens provided that liquids 1 & 2 are immiscible and chemicallycompatible, that liquid 1 is polar and conducting, and that liquid 2 isnon-conducting, which essentially implies that liquid 2 is non-polar aswell.

As those skilled in the art of chemistry will appreciate, some liquidsthat are unable to conduct electricity in their pure state will becomeconductive when relatively small amounts of appropriate substances aredissolved or mixed in as additives. For example, water and acetic acidare non-conductive in their pure state. But water becomes conductivewhen a small amount of salt is dissolved in and acetic acid becomesconductive when a small amount of water is mixed in. As those skilled inthe art of spectroscopy will appreciate, the contribution of suchadditives to transmission losses will be proportional to theirconcentration.

U.S. Pat. No. 6,694,175 teaches a use of tiny heat lamps to warm apatient. In effect, any radiation absorbed by the body and translatedinto thermal energy will warm the patient. Similarly, electromagneticradiation may be applied to the patient at various wavelengths andintensities in the form of phototherapy. However, electromagneticradiation applied to the patient will be subject to a consideration oftransmission losses (absorption by intervening media). The same is trueof electromagnetic radiation used for other purposes such asillumination or for heating the fluid incubation medium. In each case,the optics of transmission must be considered. Otherwise absorption (orscattering) by intervening media may produce unsatisfactory results.

Although not strictly necessary, using heat lamps to warm the patientadds flexibility to the thermodynamics and control of patientthermoregulation. However, a complication arises in that at somewavelengths water absorption is very strong. Referring to FIG. 60, atwavelengths near 2.95 microns, 100% absorption is observed through alayer of water only 10 microns thick. At a wavelength of 3 microns,referring to FIG. 4, infrared radiation from above (FIG. 4, infraredheat lamps 23) would heat the water of the fluid medium M rather thanreaching the patient P; referring to FIG. 54A, at wavelengths near 2.95microns even a thin layer of water 161 would be heated by infraredradiation from below before reaching the patient P; however, referringto FIG. 54B, by minimizing any intervening water, absorption by waterwill no longer be such an issue.

Those skilled in the art of spectroscopy will recognize that anemissions spectrum also indicates an absorption spectrum. Accordingly,an absorption spectrum (e.g., of the patient's body) can be collected byradiometry either by studying transmission losses or indirectly bystudying emissions. Referring to FIGS. 48A-E, different features of thebody may have different emissions spectra; in turn, this implies thatsome parts of the body absorb more strongly at certain wavelengths thanat others. By having the relevant emissions spectra in hand, orequivalently the absorption spectra, it is a simple matter to selectwavelengths absorbed by the body more so than by intervening media, soas to preferentially warm the body, or selected regions of the body, asopposed to intervening media or regions of the body not selected. Thissame principle applies to phototherapy in general. Also, an opticalfocus of heat lamps or phototherapy light sources may be employed tobetter direct electromagnetic radiation onto the patient. For example,an infrared wavelength below 1.4 microns (to minimize absorption bywater) can be used to warm the patient with the aid of optical focus. Inanother example, an absorption band of the three-layered glycoproteinmatrix that forms the shell of the human egg can be preferentially usedto heat the shell of the egg.

Infrared radiation in a region of strong water absorption (e.g., 2.95microns) can be used to heat the fluid incubation medium or other fluidsin the FCA environment. Because of strong absorption the heating will behighly localized about the area being illuminated to a depth of about 10microns or so. However, for relatively thick streams or bodies of fluid,wavelengths of moderate absorption may be preferred so that theradiation is able to penetrate deeper for more even heating. Heating maybe further localized by applying a gold surface coating, for example, toreflect the infrared radiation away from areas not to be heated.

Roithner Lasertechnik GmbH (Vienna, Austria) supplies light emittingdiodes having various wavelength ranges in the 255 nanometer to 7.0micron spectral range. Other light sources, including in combinationwith optical filters, may be used to warm the patient or to providephototherapy.

Absorption spectroscopy (transmission spectroscopy), emissionspectroscopy, scattering spectroscopy, and reflectance spectroscopy canbe used to investigate the optical, chemical, or physical properties ofa patient, fluid, or material. In an application of emissionsspectroscopy, Malchoff et al disclose a non-invasive glucose monitor forblood measurements based on thermal infrared emissions spectroscopy.Their technique employs a thermopile with a variable filter fordetecting infrared radiation from 7.7 to 14.1 microns; because glucoseabsorbs (and, thus, emits) at characteristic wavelengths (e.g., a mainband at 9.8 microns) and because the radiation amount varies withglucose concentration, glucose levels can be determined in reference tocalibration. Although absorption and reflectance spectroscopy may leadto similar results, this technique has the advantage of being purelynon-invasive. (Malchoff et al, “A Novel Noninvasive Blood GlucoseMonitor,” Diabetes Care, Vol. 25, No. 12, December 2002, pp. 2268-75)Consequently, this technique can be used to non-invasively monitorchemical levels (e.g., glucose) inside a prenidial infant's body.Similarly, spectroscopy techniques can be employed to monitor chemicallevels in the fluid incubation medium or in other fluids of the FCAenvironment.

In view of human anatomy during prenidial life, it will oftentimes bedesirable to image temperatures for different parts of the baby's bodyas a whole. For example, as the baby grows inside the egg it will bedesirable to image temperatures for tissues of the formal bodydistinctly from those of the peripheral body; it will also be desirableto image a temperature distribution within the fluid of the baby'sspacesuit. Similarly, a temperature gradient around the baby (due toheat dissipation) is of interest to microthermography. Because cellsizes for the early embryo are larger than the spatial resolution ofhigh precision infrared cameras, thermal variations within and betweencells are also of interest. Therefore, spatial homogeneity and temporalmodulation of temperature in the tissues can also be monitored by meansof radiometry.

In the prior art, Anbar (U.S. Pat. Nos. 5,961,466; 5,999,843) teachesthe use of a thermal imaging system such as a QWIP camera to detectbreast cancer by monitoring the spatial homogeneity and temporalmodulation of skin temperature related to changes in blood perfusion.Neilson et al (U.S. Pat. Nos. 6,821,787; 6,835,574; 6,991,765) teach anapparatus and methods for infrared calorimetric measurements employing athermal imaging system.

Although patient temperature detection is best suited to a non-contactthermal imaging means, e.g., as provided by an infrared camera, this isnot necessarily the case regarding a number of devices that may beassociated with an FCA. Thermistors and thermocouples may be used inthermal contact with an FCA device to monitor device temperatures. Butan added feature of an FCA providing clear optical paths to devices isthat nematic and thermochromic liquid crystals provide a means ofmicrothermography. Thermochromic liquid crystals change color across thespectrum as they change temperature over a given range. Referring toFIG. 43, these changes in color may be monitored by an optical device(e.g., via objective lens OL2) associated with the FCA. To create acolor-coded temperature map, the typically micro-encapsulated liquidcrystals are applied to a surface of a device; with the power off, thedevice is subjected to temperature increments within a given range oftemperature response for the liquid crystals. The resulting map relatesliquid crystal colors to specific temperatures, with 1 micron spatialresolution being typical. When the device is in operation, liquidcrystal colors are digitally compared to the color-coded map by computerto determine device temperatures.

Hart Scientific, Inc. is a provider of temperature calibration andmeasurement products, including incremental temperature baths.Incremental temperature steps may be taken as an aid to calibration ofFCA devices prior to patient use.

Other desirable features of a liquid crystal microthermography systeminclude uniform illumination of a region of interest, infrared-free(cool) lighting to eliminate measurement errors due to infraredabsorption, highly polarized optics to enhance image viewing andmeasurement accuracy, and software for data acquisition and imageprocessing. A number of companies, e.g., Advanced Thermal Solutions,Inc. (Norwood, Mass.), provide liquid crystal microthermographyequipment and technology of this sort. As with optical systems ingeneral, such technology may be directly incorporated into an FCA orused in a stand alone fashion.

Polarized light microscopy is used to image and measure birefringentstructures such as the three-layered shell (zona pellucida) of the egg.Tunable liquid crystal filters are preferably employed for this purpose.Tunable liquid crystal filters are also employed for multispectralimaging (to select a particular wavelength of light) and similarly forvisible color selection. Multispectral imaging is an important techniquefor visualizing tissues based on their differing responses to differentwavelengths of light. Cambridge Research & Instrumentation (CRI), Inc.(Woburn, Mass.) is a provider of liquid crystal polarized lightmicroscopy equipment and tunable liquid crystal filters.

An auto-focus feature is used to compensate for focus drift over time inthe context of time-lapse video microscopy. High definition television(HDTV) screens and other high resolution monitors are particularlyindicated for high resolution microscopy. Time-lapse video can be storedand displayed on a screen as accelerated footage to evidence very slowlyoccurring behaviors such as hatching or an embryo's turning(somersaulting) inside the egg. Confocal time-lapse video microscopy canbe used to create three-dimensional snapshots of patient developmentthat can be played back as video. This can be used, for example, tomonitor changes in cell number, morphology, and differentiation. Acomputer can be used to assign a constant reference point so that astationary view can be generated with software despite relative changesin bodily position over time, e.g., due to somersaulting. Othermicroscopy techniques may be similarly used in a time-lapse videoformat.

Those skilled in the arts of microfabrication and optical techniqueswill appreciate that a vented microcradle employing an FCA, andparticularly a side-vented microcradle with an optical path provided bya clear bottom and an open top, is amenable to any number of importantoptical techniques, including but not limited to: bright fieldmicroscopy, oblique illumination, Hoffman's modulation contrastmicroscopy, dark field microscopy, phase contrast microscopy,differential interference contrast (DIC) microscopy, fluorescencemicroscopy, confocal microscopy, deconvolution microscopy, polarizedlight microscopy, stereomicroscopy, and optical coherence tomography;infrared spectrometry (e.g., for chemical analysis), and optical Dopplertomography (e.g., for fluid velocity measurement); thermal imaging via athermal imaging system such as an InSb camera attached to a microscopeobjective, and thermal imaging via thermochromic liquid crystals viewedunder a microscope; and, phototherapy.

It may be noted that preference for a clear bottom for a side-ventedmicrocradle does not obviate the possibility of an opaque or reflectivebottom. Dedication of the bottom to optical purposes is simply onepossible use. Other uses can include support for a cell scaffold (e.g.,to support a blanket of cumulus cells), sensor arrays (e.g., an array ofelectrophysiological sensors), biological coatings, or surfacetreatments.

The invention may be operated with the aid of an eyewear viewer (orother head-mounted display system) and a control console. TheMicrooptical Corporation (Westwood, Mass.) makes wired and wirelesseyewear viewers for medical use. Control consoles are well known in theprior art of microsurgical systems. For example, Metzler (U.S. Pat. No.6,022,088) teaches an ophthamalic microsurgical system. Becauseprenidial care is operated at the microscopic level, the presentinvention contributes to the field of microsurgery and microsurgicaltechniques and is compatible with microsurgical systems and methods.

Attention is now turned to the subject of additional microfluidic designconsiderations.

In computer science, a bus is a system of addressable interconnects usedin the routing of electronic data between devices. The “bus” principlemay be extended beyond electronics. For example, in optical computingoptical data are routed. In the present case, there is a need to routefluid data or samples. In this disclosure, a fluidic bus routes fluidamounts between devices. Such amounts or quantities may be referred toas data or samples.

As a prenidial infant metabolizes resources in the fluid incubationmedium, the chemical content of the fluid changes, and it willoftentimes be desirable to detect such changes by means of an analyticdevice associated with the FCA environment. Likewise, there willoftentimes be a need to deliver fluidic treatments to the patient. Ingeneral, there is a need to rapidly route fluidic data in an FCAenvironment to enable fast response times regarding such processes.

However, given a human embryo with an outer shell diameter of 120microns, then even at a rate of flow as high as 1 micrometer per second,it would take a whole two minutes for a flowing fluid simply to travelthe length of the patient's body. Thus, if a fluid sample is to berouted from the patient to an FCA device at the same rate of flow as thefluid ventilating the patient, then it will take a long time for thefluid sample to reach such a device located elsewhere on the FCA whereit is to be analyzed. The same is true of fluid samples to be deliveredas treatments to the patient from a device located elsewhere on the FCA.Consequently, in this type of scenario response times will be very slow.

One way of solving this problem is by exchanging parcels of fluidbetween slow and fast moving channels so as to speed fluid transit anddelivery. Referring to FIGS. 61A-B, fluid from a slow flow rate channel164 can be merged with fluid in a fast flow rate channel 165, and viceversa, to speed transit of the fluid to and from a patient and variousFCA devices. Though not shown in these figures, it will be appreciatedby one skilled in the art that valves, pumps, sensors, and variousmicrofluidic features will assist in such exchanges.

Another approach to the problem is to employ digital microfluidics tobus fluid samples around in an FCA environment. Digital microfluidicsoffers advantages such as rapid transit rates of discrete fluidquantities coupled with digital control of routing. The prior art easilyhandles microliter droplets of fluid. But a microcradle 150 microns highand 500 microns square only holds about 1/27^(th) of a microliter, or37.5 nanoliters. Thus, if the flow rate of fluid in the microcradle isas much as 1 micrometer per second, it would take about an hour togenerate enough fluid to produce a one microliter sample of fluid. So,apart from a digital microfluidic system capable of busing nanoliterdroplet samples, the analog fluidic busing method illustrated in FIGS.61A-B must be relied upon to speed transit.

To make it easier to achieve nanoliter busing in a digital microfluidicsystem, a clarification of the theory of electrowetting appears needed.Specifically, I believe that a “capacitive spreading” of charges causesfluid movement. Referring to FIGS. 62A-B, charges in a polar, conductingliquid LIQ. 1 seek to spread out based on a capacitive relationship withopposite charges present in a sidewall electrode 120. As the charges inthe conducting liquid LIQ. 1 move (see arrow) so as to spread out over agreater surface area, polar molecules “piggyback” the moving charges inthe fluid via polar bonds 166, thus causing a liquid interface 167 tomove as well.

This theory helps to clarify a distinction between ordinary capillarityand electrowetting. Referring to FIGS. 62A-B, it is evident that thehydrophobic coating 118 could be replaced by a hydrophilic coating and acapacitive spreading of conductive charges would still occur. But insuch a case a spreading of polar molecules would also occurindependently of the spreading of charges, based on their affinity forthe hydrophilic coating. In contrast, by employing a hydrophobic coating118, it enables the spreading event to be turned on or off based on thepresence of charges in a capacitive relationship. Accordingly, it istheorized that electrowetting regards a spreading of charges(electrolytes) to which polar molecules are clinging in piggyback,whereas ordinary capillarity regards an independent spreading of polarmolecules.

FIG. 63 shows a classic arrangement for digital microfluidics. A dropletof fluid is made to move by turning on a voltage between a groundelectrode and a control electrode; the droplet moves in the direction ofthe leading control electrode. But a wiring problem is encountered asthe droplet size gets smaller and smaller, because the number of controlelectrodes needed will increase. In other words, the classic arrangementroutes an electrical connection for each control electrode to a remoteswitch.

FIG. 64, on the other hand, shows a prior art arrangement for digitalmicrofluidics in which the need for remote switching is eliminated.Although both arrangements employ electrowetting, this particular one ismore specifically known as opto-electrowetting because an optical beamor stylus is used to turn control electrodes on and off. Thisarrangement eliminates the need to route electrical connections for eachcontrol electrode. Instead, the control electrodes are patterned inphotoconductive silicon and an optical beam is used to switch a selectedelectrode on. To enable illumination by the optical stylus, an indiumtin oxide coating is used as an electrode along with a hydrophobiccoating of Teflon®, such that both coatings are thin enough to betransparent. An artifact of this arrangement is that an AC voltage isrequired unlike the classic electrowetting arrangement, which relies onDC voltage; however, it is contemplated that by biasing the controlelectrode circuitry DC voltages might also be used withopto-electrowetting.

At various frequencies, and especially at high voltages, AC electricalfields may have adverse implications for human health as well as forsensitive electrical or magnetic equipment. Another problem with theopto-electrowetting arrangement concerns the use of an optical stylus,which in turn must be subject to finer and finer optical controls as thesize of the control electrodes decreases, which is needed to movesmaller and smaller droplets. The costs and bulk of such opticalequipment will be prohibitive at some point as fineness increases.

The present invention prefers a distinct arrangement for digitalmicrofluidics for droplets of extremely small size. This arrangementemploys “self-scooting” circuitry according to the invention. FIGS.65A-66 illustrate the self-scooting principle. Referring to FIG. 65A,which provides a simplified top view of a bottom layer 169 containing aseries of control electrodes CE(1)-CE(N), a droplet of conducting fluid168 (e.g., aqueous) is moved along by turning the control electrodesCE(1)-CE(N) on and off in succession. FIG. 65B provides a side view,which shows the droplet 168 moving horizontally while sandwiched betweena top layer 170 containing a ground electrode GE and the bottom layer169 containing the control electrodes CE(1)-CE(N). Referring to FIG.65A, the series of control electrodes CE(1)-CE(N) forms a track for thedroplet 168 to move on; a series of liquid contact switches is formed byconducting strips S(0)-S(N), where S(0) serves as a common pole for eachof the switches and S(X) serves as the opposite pole for the Xth switchalong the track. When the droplet 168 passes over the Xth position ofthe track, the Xth switch is thrown closed by the electricalconductivity of the droplet 168. FIG. 66 illustrates exemplaryself-scooting circuitry in general, non-limiting terms; referring toFIG. 66, when the Xth switch is closed by the droplet 168, an electroniccircuit is activated such that the control electrode at position X+1 isturned on. When the droplet 168 passes the Xth location along the track,the Xth switch will open and, using relevant circuitry, the electrode isturned off.

The circuitry shown in FIGS. 65A-66 is merely exemplary. The distinctprinciple of self-scooting circuitry for a digital microfluidic systemis that a series of liquid contact switches are employed to detect aposition of an electrically conducting droplet along a track formed bycontrol electrodes, such that an opening and closing of such switchescan be used to control a turning on and off of selected controlelectrodes, so that the droplet in effect “scoots” itself along thetrack. The droplet will move along the track until a break isencountered.

FIG. 67A shows an exemplary CMOS circuit employing NAND gates to controlthe Xth control electrode CE(X) such that droplet movement in the rightor left direction may be selectively controlled. FIG. 67C shows anequivalent circuit employing AND and OR gates. FIG. 67D shows anequivalent circuit employing tri-state buffers. FIG. 67B shows anexemplary CMOS circuit to enable or disable signals from liquid dropletcontact switches (switching electrodes) to a control electrode; forexample, such a circuit can be used to stop droplet flow along a track;similarly, referring to FIG. 67E, such a circuit can be used to stopflow along one track and to route flow onto another track. FIG. 67Eshows enabling conditions (0, disabled; 1, enabled) for controlelectrodes, such that a droplet's path is routed at a fork in the road.Referring to FIG. 67F, droplet switches can be used to enable encoding,control, or event circuitry; for example, the position of a droplet maybe encoded, a device for sensing a condition of the droplet may becontrolled so as to turn the device on, or an event may be commenced inresponse to a signal from a droplet switch.

Computer control of control electrode circuitry enables dropletmovements to be digitally controlled in response to events, sensedconditions, or software. Computer control of circuitry also enablesdroplets to be sensed, monitored, and acted upon; it also enablesdroplet events to trigger or moderate other events. Note that eachcontrol electrode or droplet position does not need to have the samecircuitry associated with it; instead, circuitry, including computercontrols, is required only as needed to handle droplets at given pointsalong a track.

Referring to FIG. 65B, simple transistor circuits, more complex CMOScircuits, and other circuitry may be directly integrated into a layer169 containing the control electrodes. However, unlike theopto-electronic arrangement of FIG. 64, control electrode electronicsmay also be routed to remotely located circuitry according to theself-scooting arrangement. For example, though remote location isgenerally not preferred, in some cases it may be desirable in specificlocations to enable a clear optical path through a droplet. This can beachieved by employing indium tin oxide coatings for electrodes andTeflon® for the hydrophilic coatings according to the art. In contrast,the optical path would be effectively blocked by circuitry placedimmediately underneath the control electrodes, as semiconductors areopaque to visible light.

Referring to FIG. 68, under the action of electrowetting in anelectrowetting region 171 a liquid 172 flows continuously from areservoir 173; in this case a series of control electrodes is notrequired for flow. But when discrete quantities of liquid such as adroplet are used, control electrodes at the trailing end of the dropletmust be turned off in order to create a preference for motion in a givendirection; otherwise the droplet will not move. For this reason, aseries of control electrodes is required to move droplets byelectrowetting. Referring to FIG. 69A, one alternative is to turn aleading control electrode on and then off again as a droplet passes;this is consistent with the exemplary circuitry shown in FIGS. 67A, C-D.Referring to FIG. 69B, another alternative, called a one-shot approach,is to first turn all control electrodes on and to then turn trailingelectrodes off to move the droplet; then at some point in time thecontrol electrodes may be reset again to the on position. Referring toFIG. 69C, control electrodes are reset essentially as soon as thedroplet has passed.

In this disclosure, two categories of electrowetting are distinguished.The electrowetting arrangement of FIGS. 29A-B relies on what is here istermed “charged” electrowetting. Referring to FIG. 29B, an electrode 119placed in contact with a conducting liquid 116 charges the liquid 116,in this case with a net positive charge. Though the charge on the liquid116 is balanced by an equal and opposite charge on an external sidewallelectrode 120 in a capacitive relationship, so that the system as awhole is neutral in charge, nevertheless the liquid 116 itself carries anet charge. Hence, this is called charged electrowetting. In contrast,FIGS. 70A-B illustrate what is here termed “dielectric” electrowetting.A droplet 174 is sandwiched between top 175 and bottom 176 electrodesthat are coated with insulative 122 and hydrophobic 118 coatings.Referring to FIG. 70A, the hydrophobic coating 118 is unwetted. However,referring to FIG. 70B, when an electric potential is applied betweenelectrodes 175-176, an electric field

is set up between the electrodes 175-176. In this example, the droplet174 contains ionic charges (electrolytes) that separate according tocharge polarity in response to the electric field

(note that there is no net charge on the droplet 174, unlike the case ofcharged electrowetting). Then, according to the principle ofelectrowetting, a capacitive spreading of the charges, coupled withpiggybacking by polar molecules in the droplet 174, will cause thecoating surface 118 to be wetted despite being hydrophobic.

Referring to FIGS. 70A-B, note that although a droplet containing mobileionic species is preferred to enable better charge separation andmigration to opposite electrodes, in theory even a non-conducting polardroplet will exhibit dielectric electrowetting if the electric field

is strong enough to cause polar molecules to orient and spread accordingto their dipole moments. Referring to FIGS. 70A-B, note that for adroplet containing mobile charges the ability to separate charges ofopposite polarity is limited by the dielectric constant of the dropletand the strength of the applied electric field

. Note also an opposing relationship of electrodes is required fordielectric electrowetting in order to set up an electric field

. From a physical perspective, this is the same relationship as that ofa parallel plate capacitor, such that the dielectric medium between theplates happens to be an electrolyte solution that is insulated fromelectrodes. In contrast, charged electrowetting derives its chargestoring power from the principle of an electrolytic capacitor.

Referring to FIG. 29B, a net charge in a conducting liquid 116 iscreated by a voltage potential applied to the liquid 116 in the mannerof an electrolytic capacitor. Because electrolytic capacitors can storemore charge energy than parallel plate capacitors, in general it iscontemplated that charged electrowetting will offer more power to movefluid in a microfluidic system than dielectric electrowetting.

Referring to FIG. 71, the dielectric electrowetting arrangement of FIG.65B could be practiced as a charged electrowetting arrangement instead.For example, referring to FIG. 65A, electrode S(0) could be employed tocharge the droplet 168 with a charge opposite to that placed on controlelectrodes CE(1)—CE(N). Note that no ground electrode (GE) is requiredon top, in contrast to the case of dielectric electrowetting. Instead,in this case a top plate 177 simply serves to flatten the droplet 168 toincrease surface contact with the bottom plate 169 while the hydrophobiccoating 118 helps to reduce friction. Notably, remote switching, opticalswitching, and self-scooting circuitry are all compatible with the FIG.71 arrangement.

Referring to FIGS. 63-64 of the prior art, as well as FIG. 65B accordingto the present invention, note that the top plate (ground) electrodedoes not participate substantially in capacitive spreading, given thatcharge is distributed in the ground electrode more or less equally onboth sides of the droplet, in contrast to the control electrodes, whichlead the droplet in a given direction by turning leading controlelectrodes on and trailing ones off. In contrast, referring to FIG. 72,a series of ground electrodes GE(1)-GE(N) in a top layer 178 can beturned on and off at the same time as the control electrodesCE(1)-CE(N), which have opposite polarity. That this dielectricelectrowetting arrangement is more powerful than the others is indicatedby a singly-wetted meniscus 179 in the FIGS. 64 & 65B arrangementsversus a doubly-wetted meniscus 180 in the FIG. 72 arrangement.

FIG. 73 shows a double-sided, charged electrowetting arrangement. Inthis case, control electrodes CE(1)-CE(N) face each other in the top 178and bottom 169 and an electrode in contact with the droplet 168 (e.g.,electrode S(0) shown in FIG. 65A) is used to charge the droplet with anopposite charge. Although the FIG. 73 embodiment could conceivably bepracticed in an annular embodiment, to manufacture it would bechallenging for small sizes of the control electrodes. In contrast,embodiments with electrodes placed at, say, 90 degrees to each other,though more challenging than a top and bottom layer arrangement, wouldnevertheless appear to be more feasible by comparison. Theelectrowetting arrangements of FIGS. 72-73 can be practiced either asremote switching or self-scooting embodiments; however, in general, useof a semiconductor in both layers 169, 178 will render the layers opaqueto visible light, and so optical control of electrodes (i.e.,opto-electrowetting) will not be possible, unless, for example,photoconductivity can be established in a semiconductor layer by meansof a wavelength of light to which the semiconductor is transparent,e.g., infrared light.

FIG. 74 shows an open-top charged electrowetting arrangement for digitalmicrofluidics; an exposed electrode S(0) (not shown) maintains a chargein a moving droplet that is opposite to a charge in the controlelectrodes C1-C(N) when turned on. Remote switching, optical switching,or self-scooting circuitry can be employed with this arrangement. Aswith other arrangements, such an arrangement can be practiced using anydesired pattern of electrodes (e.g., a two-dimensional planar array ofcontrol electrodes). However, consider a surface coating 181. If thesurface coating is hydrophilic, the bottom edges of a droplet will bemade to spread out on the floor; this places the leading edge of thedroplet in contact near the control electrodes; but even though thiscontact improves the force exerted on the droplet by a controlelectrode, this force will be counteracted by frictional forces due tothe hydrophilic nature of the coating; however, as droplet size getssmaller, these frictional forces will be proportionately less comparedto the force exerted on the leading edge of the droplet. In contrast, ahydrophobic or superhydrophobic coating will make the droplet form aball and so there will be less contact between the leading edge of thedroplet and the surface where control electrodes are embeddedunderneath. However, coulombic forces will exert more influence asdroplets get smaller (see FIG. 75).

In general, the switching electrodes carry some charge and so thepolarity and voltage of the switching electrodes becomes an issue inreference to their location inasmuch as they may partially blockcoulombic interaction with control or ground electrodes. Also, thehydrophilic versus hydrophobic quality of an electrode such as aswitching electrode or an electrode used to charge a droplet becomes anissue in reference to its contact with the droplet. Although a patternof switching electrodes S(0)-S(N) in FIG. 65A is exemplary, electrodesmay be placed and routed electronically in any desirable fashion. Forexample, referring to FIG. 71, electrode S(0) may be placed on the toplayer 177 and S(1)-S(N) may be placed over and routed through controlelectrodes CE(1)-CE(N) on the bottom layer 169 using vias that connectto self-scooting circuitry in the bottom layer 169, which is formed of asemiconductor, and S(0) may be wired down to the circuitry in the bottomlayer 169. In another example, electrode S(0) may be placed side-by-sidewith electrodes S(1)-S(N) on top of the control electrodes CE(1)-CE(N)and routed to circuitry underneath.

In general, control electrodes may take on any variety of shapes; forexample, electrodes may be shaped to match the leading edge of adroplet. In cases where bi-directional droplet transit is desired andelectrodes have an asymmetric shape, two layers of control electrodes,formed to serve respective transit directions, may be placed one on topof the other and separated by an insulating layer. Various stencilpatterns may be used to guide droplets in layers. However, such patternsdo not necessarily need to provide lateral support for a droplet, unlikethe case of analog microfluidics. Borders may be used to guide dropletsin open-top arrangements, including borders made of a contrast ofhydrophilic and hydrophobic coatings.

Referring to FIG. 30B, a liquid lens according to the invention willmove under the action of charged electrowetting in the manner ofcapillarity unless valves controlling either microfluidic channels 128or 129 are closed to prevent fluid transit. With the valves closed, thelens will change shape rather than moving as a whole.

In general, self-scooting circuitry will require a semiconductor layerin which switching operations are performed by integrated circuitry.However, a special exception occurs. In this simple case, calledtransistorless self-scooting circuitry, the circuitry at minimumrequires only conductive elements. Of main interest is a dielectricelectrowetting arrangement. Referring to FIGS. 76A-B, control electrodesCE(0)-CE(N) are patterned on a bottom layer 182 and each controlelectrode CE(0)-CE(N) is separately connected to its own electricalcontact pad 184 by a conductive trace 183. A top layer 185 contains aground electrode GE. FIG. 76A shows the bottom face of the top layer 185and FIG. 76B shows the top face of the bottom layer 182. The top layer185 is aligned over the bottom layer 182 as indicated by an alignmentmarking ▴ such that the ground electrode GE goes over the controlelectrodes CE(0)-CE(N). Not shown is a middle layer to provideseparation between the top 185 and bottom 182 layers. Referring to FIGS.77A-B, the electrode surfaces of both layers 182, 185 are coated with ahydrophobic layer 186, which also needs to be electrically insulating onthe bottom layer 182.

However, on the bottom layer 182 the contact pads 184 are left exposedand a switching contact region S(1)-S(N) is left exposed on respectivecontrol electrodes CE(0)-CE(N−1). The control electrodes CE(0)-CE(N) areinterdigitated such that the switching contact region S(X) associatedwith control electrode CE(X−1) extends in the manner of a digit into aninvaginated region of control electrode CE(X). If a hydrophobic orotherwise relatively frictionless electrode material is available, thenthe top layer 185 does not necessarily need to be coated. Otherwise,assuming a hydrophobic coating 186 that is also electrically insulating,a series of holes or a continuous line can be patterned in the coating186 so that a switching electrode S(0) is exposed while at the same timeminimizing droplet friction with the ground electrode GE.

FIG. 78 is a side cross-sectional view taken along a line 187 withrespect to FIGS. 77A-B. According to operation, electrical contact (notshown) is made between the ground electrode GE and each of the contactpads 184 so as to charge the ground GE and control electrodesCE(0)-CE(N) in the manner of parallel plate capacitors. The charging canbe accomplished, for example, by drawing a brush-type electrical contactacross each of the contact pads 184 or by employing a clamp-type contactswitch to contact each of the pads 184. Once the electrodes GE,CE(0)-CE(N) are charged, all of the control electrodes CE(0)-CE(N) areturned on, which is the time t0 condition shown in FIG. 69B for aone-shot arrangement.

Referring to FIG. 78, a droplet 188 formed of a conducting liquid isshown to move as a trailing control electrode CE(2) is turned off byelectrical discharge 189 as an electrical contact is made by the droplet188 between switching electrodes S(0) and S(3). In other words, thedroplet 188 is made to move as trailing control electrodes are turnedoff via electrical discharge through the droplet 188 itself.

FIG. 79A shows a basic circuit diagram for transistorless self-scootingcircuitry. This circuit corresponds to the one-shot operation shown inFIG. 69B, except that discharge does not occur as a square wave inasmuchas the droplet 188 has some electrical resistance. FIG. 79B shows aself-refreshing circuit diagram for transistorless self-scootingcircuitry that follows the pattern shown in FIG. 69C, such that trailingelectrodes are refreshed after they are turned off, except that theresetting does not occur as a square wave due to a resistor R; theresistor, which should have a resistance that is larger than theresistance of the droplet 188, enables a control electrode to be turnedoff momentarily as the droplet 188 passes, but the control electrodeturns on again after the droplet 188 has passed.

To practice transistorless self-scooting circuitry in a chargedelectrowetting arrangement, a net static charge has to be placed oncontrol electrodes. An opposite charge on the droplet is maintained byan electrode S(0) while discharge is enabled via switching electrodesS(1)-S(N) so as to turn trailing electrodes off. However, whether placedon droplets or on electrodes, net static charges will generally not bepreferred.

Sometimes droplets or their contents will not be tolerant of an electriccurrent passed through them in any way. For example, a droplet maycontain chemicals that may react electrochemically, or the droplet maycontain a cell or virus that is being manipulated by digitalmicrofluidics. In such a case, referring to FIG. 80, a parallel or“pacer track” circuitry may be employed. In this case, a pacer track 190controls movement of a pacer droplet 168. The movement of the pacerdroplet 168 controls underlying circuitry associated with the turning onand off of control electrodes associated with the pacer track 190 aswell as those associated with a “free track” 191 containing a freedroplet 192. FIG. 81 shows an exemplary circuit diagram for a pacertrack arrangement, which is related to the circuit diagram of FIG. 66.Note that the free droplet's 192 movement is controlled by the pacerdroplet's 168 movement.

The free droplet 192 does not need to be subject to currents associatedwith the switching electrodes S(0)-S(N), since those currents are borneby the pacer droplet 168. However, in a charged electrowettingarrangement, the free track 191 will still need to maintain an electrodein continuous contact with the free droplet 192 so as to maintain a netcharge in the manner of an electrolytic capacitor, as previouslydescribed. However, in an electrically well-insulated system, a netstatic charge might be maintained without such an electrode.

Referring to FIG. 78, surface features on surfaces where the droplet 188travels become important, especially as electrode size gets smaller andsmaller. In this example, the surface is figuratively shown to besomewhat planarized, with the switching electrodes S(0)-S(N) beingsomewhat raised to form slight nubs. The importance of surface featuresis evident in that the passing droplet 188 must be able to makeelectrical contact with the switching electrodes S(0)-S(N). At the sametime, it is desirable to reduce friction.

Referring to FIG. 82, although an interdigitated arrangement of controland switching electrode contact regions has been shown in FIG. 76B,other arrangements are also possible, including by use of vias. Also,discharge currents may be used to drive other circuitry. Referring toFIG. 83A, in some cases two or more different tracks of controlelectrodes 193-194 may be separated from each other by insulation, sothat, for example, droplet travel in respective directions can becontrolled by respective tracks; the control electrodes of differentlayers may share circuitry to some extent (e.g., a ground electrode).Referring to FIG. 83B, at selected locations having two or more layersof separately controlled control electrodes 193-194 may help to preventdroplets from getting stuck at points. Referring to FIG. 84,interdigitated control electrodes are known in the prior art to permitclose contact. However, as control electrodes become very small, it maybe preferable to slightly overlap control electrodes in a given controlelectrode layer 195; although FIG. 85 shows overlapping in a staggeredfashion, a fallen domino pattern, though challenging to manufacture, mayalso be considered.

Referring to FIGS. 29A-B, in the absence of a hydrophilic surface, awater droplet 116 wants to revert to a droplet shape, as shown in FIG.29A, based on its own surface tension (mutual attraction of polarmolecules). However, a shape of the water droplet 116 as shown in FIG.29B will be obtained as the forces of surface tension arecounterbalanced by the forces of capacitive spreading as shown in FIGS.62A-B. Either polarity may be used since either way the charges willparticipate in capacitive spreading. Notably, since the charges aremaintained by capacitance, the shape of the water droplet 116 will bemaintained even when disconnected from a battery that established avoltage potential difference V. Stronger voltages will overcome agreater amount of surface tension, and so the shape of the water droplet116 will change more. However, without an insulating layer of oil 117charges will jump from the water droplet 116 and spread by themselvesover an opposing sidewall electrode 120 if the voltage is high enough.In turn, this neutralizes the effect of the potential difference.Referring to FIG. 70B, the same sort of thing will happen even in thecase of dielectric electrowetting. As droplet size gets smaller andsmaller, and as microfluidic power is maximized by using an annulararrangement as shown in FIG. 30B or by using an arrangement such asshown in FIGS. 72 & 73 as compared to an arrangement with less power asshown in FIG. 65B, the amount of voltage required to move or manipulatea droplet will be lowered. If low enough, use of an insulating fillerfluid will not be essential. However, for larger droplets or lessefficient arrangements, or even for small droplets, it is contemplatedthat in the future nanotechnology will someday provide a means to switcha surface back and forth between hydrophobic and hydrophilic states. Insuch a case, control electrodes will be replaced by “control surfaces”;yet, the use of self-scooting circuitry to switch the control surfaceson and off will be equally pertinent. Of note, the paper by Rosario etal (Ibid.) provides a primitive example of turning a control surface onand off in response to light so as to move a droplet.

One skilled in the art of digital microfluidics will appreciate thatself-scooting circuitry according to the invention will enable dropletevents such as dispensing, merging, mixing, rotating, injecting,locating, sensing, conditioning, moving, routing, and so on to bechoreographed with much greater control based on an ability to finelydetect droplet position electronically. It will also be appreciated thatself-scooting circuitry will enable a more intimate control of dropletsand a control of finer droplets than has been previously realized.

Attention is now turned to the subject of additional designconsiderations concerning incubator environmental controls.

In the prior art, human embryos are kept in incubator ovens and aremoved back and forth manually in laboratory dishes between the oven anda microscope stage for examination and treatment. This introducesnumerous problems, such as poor control of temperature and handlingerrors such as spillage. Whether inside a humidity controlled oven oroutside the oven on the microscope, evaporation of water from the fluidincubation medium means the concentrations of dissolved substances(osmolarity) will change over time due to water loss. Alternateapproaches have included containing the embryo within a completeenclosure such as a tank-like chamber (Thompson et al, U.S. Pat. No.6,673,008; Campbell et al, US published application 2002/0068358) or amicrochannel (Beebe et al, U.S. Pat. No. 6,193,647); though theseapproaches serve to reduce evaporative losses, complications abound.

For one thing, unlike the open-top arrangement provided by the presentinvention, patient access is hindered. Also, ambient pressure is notindependent of a pressure used to urge fluid to flow past the embryo insuch devices; and, as anyone familiar with scuba diving knows, osmoticbalance in the tissues is related to fluid pressure; thus, with someirony, in some cases due to pressure variations, osmotic balance mightbe upset more so than it would have if osmolarity had even been allowedto vary slightly.

However, these complications are unnecessary. For one thing, in aflowing system it is an easy matter to correct for evaporative losses asthey occur. For example, referring to FIG. 6D, exemplary FCA device D1might be an osmolarity compensator. The purpose of the osmolaritycompensator (or regulator), then, is to correct for changes in watercontent in the fluid incubation medium M, as caused, for example, byevaporation.

For another thing, evaporative losses can be substantially controlledand prevented in the first place by means of a number of environmentalcontrols. For example, a laminar and substantially lateral flow of airfrom an air system located intimately above a vented microcradle's opentop can be combined with a substantially vertical flow in the form of alaminar flow hood. The function of the lateral flow is to maintainparameters of air temperature, humidity, and gaseous content (e.g., CO₂content) in proximity to the vented microcradle, whereas the function ofthe vertical flow is to maintain temperature and to remove stray amountsof humidity and air at the given composition that are not desirable foroperators.

Referring to FIG. 86, lateral 196 and vertical 197 airflow systems haverespective inlets 198-199 and outlets 200-201 for the entry and exit ofair. In other words, the flow systems function in the manner of aircurtains, and not merely in the sense of a laminar flow system thatprovides only for air suction. As shown in FIG. 86, the lateral flowsystem 196 is intimately located above a vented microcradle 202 and mayeven be integrated with an FCA 203 containing the microcradle 202.

In practice the need for lateral airflow as shown in FIG. 86 willgenerally be limited to a subtle amount of flow, being that the fluidincubation medium bathing the patient will most likely be discardedafter passing by over the patient for a short period of time, includingin a recirculating or to-an-fro mode of fluidic ventilation. Thus, therewill generally not be a great deal of time for evaporation to occur. Thediscarded fluid should be kept for scientific analysis in reference topatient outcomes and in reference to data collected for the patientduring the time in which the given quantity of fluid was used.

Referring to FIG. 6D, exemplary FCA device D3 might be a device fordiscarding used fluid M and replacing it with fresh media M, includingsequential media or media selected according to patient needs.Similarly, although various devices can be used to maintain the gaseousand other content parameters of the fluid incubation medium M, ingeneral the medium M will simply be refreshed with new medium M asneeded.

As an alternative or complement to use of a lateral airflow system 196as shown in FIG. 86, referring to FIG. 87 microfluidics can be employedto provide a horizontal laminar flow curtain 204 in an upper portion ofa vented microcradle 205 to isolate a ventilation region 206 below fromthe environment above the open top 2 of the microcradle 205. The laminarflow curtain 204 contains substantially the same fluid content of theincubation medium M as is needed for patient health. Based on laminarflow, which is a characteristic of microfluidic systems, the curtain 204provides isolation to protect the ventilation region 206 from exposureto the environment. In other words, the flow curtain 204 experiences theeffects of exposure to the environment above, rather than theventilation region 206 having to.

In special cases, an FCA may be provided with a bell jar type coverhaving air inlets and outlets to provide environmental isolation, forexample, in a portable embodiment. In general, a MEMS-actuated cover maybe provided on top of an FCA to cover the open top of a ventedmicrocradle for whenever added protection or temporary isolation isneeded. For example, an earthquake sensor might trigger the actuation ofthe cover to protect the patient from spillage during an earthquake. TheMEMS-actuated cover is preferably made of a covering material that istransparent to light at suitable wavelengths. By MEMS-actuated is meantactuated according to micro-electro-mechanical systems technology(MEMS).

As a stationary incubator platform for the care of human infants, theinvention is best operated in an industry standard clean roomenvironment. Clean room technology is well known to the semiconductorindustry to keep dust particles from falling on semiconductor chipsduring manufacture. In the prior art, practitioners of in vitrofertilization have generally relied on laminar flow hoods to keep dustparticles from falling into a dish in which an embryo is kept. However,although a flow hood 207 as shown in FIG. 86 can be used to an extent inthis fashion, exclusive reliance on a flow hood to keep particles awayfrom the baby may be inadequate. In contrast, industry standard cleanroom practices may be confidently relied upon, as these are wellestablished to ensure a particle free environment. Those entering theclean room environment should be free of volatile substances andparticulates, and gowns and face coverings should be worn at all times.The design and construction of the clean room should be free of volatilesubstances and other contaminates (e.g., residues from constructionadhesives) with special emphasis on teratogenic or other toxicsubstances. A separate fume hood or ventilated storage area should beprovided for storage of volatile or potentially hazardous articleswithin the clean room. Sliding glass doors, preferably verticallysliding or laterally folding, and preferably double-walled, may beemployed about the patient flow hood 207 to provide added isolation forthe patient when operators do not require access.

Pressure, temperature, and humidity in the clean room environment areclosely regulated. However, the temperature and humidity, along with aircomposition, are preferably set in a manner suitable for operatorcomforts. Clean room air pressure should be the same as ambient pressurefor patients. Care should be taken so that ambient pressure is notdisturbed when sliding glass doors are shut around the patient flow hood207.

FIG. 88 shows an exemplary layout for a fertility intensive care unit(FICU) clean room containing three micro intensive care units (MICUs).Each MICU contains a prenidial incubator (a) on a table platform with aflow hood on top. The flow hoods are preferably supported from theceiling so that when vertical sliding glass doors are lifted the entiretable platform is left unobstructed by vertical supports. Accordingly,the sliding glass doors have interlocking corners that permit slidingagainst each other. Nurses' stations, patient transfer rooms, patientrooms, operating rooms, family rooms, and other usual medical facilitiesare provided outside the FICU clean room environment.

Prior to each use, prenidial incubators and their environments should beclean, sterile, and non-pyrogenic. They should also be free ofendotoxins, teratogens, and other toxins.

The invention requires intensive care medical operation at the microlevel. A number of existing medical disciplines may contribute to suchoperation. However, of the existing medical disciplines, microsurgeryand intensive care nursing provide the most closely related medicalfields to operate a prenidial incubator according to the invention.Obstetrics and obstetrical nursing also provide closely related medicalfields in view of an ability to ease the transition between prenidialincubation and patient transfer. Gynecology, endocrinology, and a numberof subspecialties of biology also provide relevant expertise.

Referring to FIG. 48E, human hatchlings are especially sensitivebecause, unlike embryos inside their protective egg capsules, thetissues of hatchlings are directly exposed. Also, because hatchlings arededicated to implantation for survival, contact with an artificial(non-implanting) surface can be hazardous and unhealthy. For example,they may attempt implantation on an inappropriate surface of a prenidialincubator. Referring to an exemplary FIG. 8 of the parent application(Ser. No. 10/908,861), fluid flow in the upward direction through thevented flooring can be used to lift the patient above the flooring. Itis therefore possible to retain the patient in a fixed location within acolumn of fluid flow, based on the same principle of Bernoulli'sequations in fluid mechanics that enables a ping pong ball to levitateover a column of air. In this way, the patient will not contact anysurfaces during incubation. Using fluid pressure variations or relevantorientations of fluid streams, it is further possible to rotate thepatient.

The use of water effects to provide a massaging action is well known. Itis therefore also possible to provide a therapeutic water massage forhatchlings using streams or fluid pressure variations. Because prenidialinfants are kinesthetically active, it is contemplated that watermassage may offer a useful therapeutic benefit, particularly forhatchlings. In the maternal body, a massaging effect is likely to beprovided by maternal movements, contractions, and the pulsing of thematernal heart and circulatory system.

Referring to FIGS. 89A-D, the vented flooring 208 for a microcradle mayinclude any desirable pattern of openings for fluid. In FIG. 89D, araised feature from a layer below (here shown in the exemplary shape ofa plus sign) is provided as a breach guard. To rely on a floor opening(as opposed to a side opening) to provide rotary fluid flow, openingsfor fluid may be etched at an angle to the vertical to provide rotaryflow.

Referring to FIG. 90, a side-vented microcradle 209 can be employed tolift a patient P off the bottom 3 by urging fluid to flow in an upwardpattern 210 via side vents (ventilation ports) at a rate needed to liftthe patient P, as illustrated. Additional ports for fluid flow (from thebottom, sides, or above the microcradle 209) may be used to producevarious flow effects. To provide a lifting of the patient P, the patternof fluid flow 210 may preferably be a vortex pattern caused by anglingfluid streams in a vortex pattern.

The invention provides for a control of fluid flow and other parametersof the care environment based on feedback from patient temperature andother indicators of patient health status. Other indicia (e.g., chemicalcontent of flow samples) also provide feedback.

Devices are contemplated to enable biofeedback between a mother and herexternally incubated child, including by means of wirelesscommunication. For example, a device placed inside or in fluidiccommunication with the mother may release an amount of human chorionicgonadotropin into her fallopian tube, uterus, or other part of her bodyin response to production of the same by an infant in a prenidialincubator. Biofeedback of this sort will enable mother and child to“synchronize” in preparation for implantation. In another example, atransfer catheter may be equilibrated with uterine temperature. Inanother example, fluid flow ventilating the patient in a prenidialincubator may be made to pulse in response to indications of thematernal heart rate. Careful examination of parameters and conditions ofnatural gestation will lead to a discovery of parameters and conditions(e.g., the infrared output of the maternal body inside the womb) thatare suitable for engineered provisions as well. In general, parametersassociated with the invention or its operation may be refined based onknowledge of the human body.

Prenidial incubators can be designed to accommodate different patientparameters. For example, human eggs are naturally covered with coronaradiata cells. These cells, which are maternal cells surrounding theshell of the egg, have been traditionally removed prior tofertilization, leaving the surface of the egg bare. However, techniquesto incubate patients with the corona radiata cells intact, or intact ona region or hemisphere of the egg, will generally require a larger sizeof the incubator due to a larger effective size of the egg. Similarly,if hatchlings are to be accommodated then the incubator's size willgenerally have to be larger than for embryos alone; also, the infantmust be protected from attaching to or breaching a structure of theincubator (or transfer catheter). In general, it is contemplated thatresearch into a use of structures with breachable apertures to enable ahatchling to repeat his or her hatching behavior for exercise, researchinto a use of hydrotherapy, and research into a patient's behavioralsensitivity and response to chemical, optical, surfacial,electrophyiological, biologicial, mechanical, and physical stimuli maybe explored. In other words, a variety of therapeutic novelties may beconsidered in the context of the controlled care environment provided bya prenidial incubator.

Referring to FIG. 91, Cecchi et al employ a picket fence structure forthe dual purpose of enclosing embryos and for stabilizing a microdrop offluid incubation medium under which the embryos are submerged. However,as illustrated in FIG. 91, a problem arises in that the pickets are notarranged in the shape of the drop, leading to a difference instabilizing resistance depending on whether the drop is accidentallyaccelerated (e.g., by tilting under gravity when dishes containingembryos are moved) in a horizontal versus diagonal direction. Anotherproblem is that the pickets, as a stabilizing structure, are notarranged substantially near the circumference of the droplet. Theproblem with this is that the drop will be allowed to acceleratesubstantially before encountering any stabilizing resistance.

In general, when practiced as a stationary incubator platform a problemof tilting under gravity is not a major issue with respect to thepresent invention, because there is no need to move patients back andforth between incubator ovens and microscope stages as there is with theart of Cecchi et al. At any rate, referring to FIGS. 92A-C, the presentinvention prefers to use stabilizing structures for a droplet 211 thattake the same shape as the circular circumference of the droplet 211 andare placed substantially next to the droplet's 211 circumference.Referring to FIG. 92A, to stabilize the droplet 211 in which a ventedmicrocradle 212 is submerged, an independent set of pickets 213 forms acircumferential structure just inside the droplet's 211 circumference.Pickets may also be elongated to form standing plates along radial linesemanating from the droplet's 211 center. Shaping the outer verticalcontours of the pickets to match the contours of the droplet 211 willmaximize resistance. Stabilizing pickets or plates placed on the outsideof the droplet 211 should be hydrophobic, whereas those placed on theinside may be hydrophilic or hydrophobic (since either way the droplet211 will resist breaking its surface tension) but are preferablyhydrophilic. Referring to FIG. 92B, alternatively, a hydrophilic channel214 may be etched just inside the circumference of the droplet 211. Ifthe surface on which the droplet 211 rests is hydrophobic, a hydrophilicstrip may be used on the inside surface in place of a channel 214.Alternating concentric hydrophobic and hydrophilic strips (not shown)may also be used adjacent to the droplet's circumference. Also, if theinner surface on which the droplet 211 rests is hydrophilic ahydrophobic area may be placed around the droplet's 211 outercircumference. Referring to FIG. 92C, preferred is a stabilizing wall215 on the outside of the droplet 211.

Generally speaking, although a vented microcradle is provided withrelatively small ports for fluid entry and exit, one or more largerholes, or alternatively high velocity channels, may be provided for thesake of quick refilling or refreshing of the fluid in the microcradle.For example, patient health status, as indicated by monitors, maywarrant an emergency treatment with an alternate fluid or medicine.Ports introducing medications or other fluidic treatments can bearranged to improve mixing in the microcradle to ensure proper patientexposure. Fluorescent dyes and other techniques can be used to assessmicrocradle hydrodynamics (e.g., mixing) when contemplating newconfigurations for a vented microcradle. In the drawing, a figure suchas FIG. 7B shows a via 41 forming walls of a microcradle provided with atotal of eight ventilation ports 4 at the bottom of the cradle arrangedin a given spacing arrangement. But in general such ports may beprovided in any number as needed, with nominal spacing or even under acontinuous shelf.

Metals such as copper, if leached into the fluid of the patient's fluidincubation medium, would be highly toxic. For example, a structuralfailure, such as a leakage of fluid between FCA layers due to imperfectbonding, might enable metal-fluid interaction in places where suchcontact is normally impossible. In contrast, employing metals such asgold or platinum in such places would be safer. Safety coatings may alsobe applied to ensure the chemical and electrical inertness of underlyingmaterials in relevant areas of potential hazard.

When a microdrop configuration is employed, problems with fluid flowinto and out of the microcradle may result in an imbalance that causessignificant changes in the volume of fluid bathing the patient. To avoidthis potentially disastrous situation, an alarm should be provided todetect changes in the fluid volume of the microdrop. Such changes canalso provide feedback for the control of fluid flow events. Referring toFIG. 93, an optical alarm or volume-sensing device employs a beam oflight 216 from a light source directed through the meniscus 217A of thedroplet in a lens-like fashion, which is then detected at a point P_(a)on a photodetector array 218; changes in droplet volume are registeredby the photodetector 218 based on corresponding changes in the shape ofthe meniscus 217B, which cause the light 216 to be focused on adifferent point P_(b). In general, the beam of light may traverse thedroplet in any number of configurations, including downward (as shown),upward, or sideways through the droplet to a photodetector, or the beamof light may be reflected back to a photodetector after passing throughthe meniscus. The light source should provide a beam of light at awavelength or wavelengths that do not heat or otherwise affect the fluidof the droplet or FCA. An optical means of detecting volume changes hasthe advantage that no electricity is used in contact with the fluid ofthe droplet, unlike use of an electrical sensor. As an alternative orcomplementary technique, changes in droplet size can be monitored bymeans of recording a digital image of the droplet and processing it by acomputer.

Specific heat capacity is a measure of the amount of heat needed toraise the temperature of an amount of substance by a given number ofdegrees. Low specific heat capacity means small changes in heat transferwill produce large changes in temperature. Generally, a fluid with highspecific heat capacity, such as water, will be preferred for the runningfluid of the temperature bath because large heat transfer is possiblewithout changing the temperature much. However, in some cases it mayalso be desirable to heat or cool a branch of running fluid quickly to acertain temperature for the benefit of an auxiliary device, such as amicro chem lab, that requires a certain temperature for a brief periodof operation, or a spectrum of temperatures over a given period of time.In such a case, a running fluid with a low specific heat capacity wouldbe desirable. Separate baths may also be provided for this purpose.

In general, it is desirable to minimize electrical effects in aprenidial incubator because, for example, either the patient orsensitive electrophysiological equipment may be adversely affected.Faraday cages, including ones built into the FCA, may be provided toimprove electrical isolation. Indium tin oxide may be used as a faradaycage material wherever optical transparency is needed. Reliance onpneumatic and mechanical components over electrical ones can also helpto reduce electrical effects.

Although each FCA is generally dedicated for single use only, a numberof FCA components can be recycled after each use to help lower costs.

Microfabrica, Inc. (Van Nuys, Calif.) has developed a microfabricationplatform called EFAB™ for the fabrication of 3-dimensional microdevices.Presently, the technology only works with metals. However, if a similarmethod could be developed for use with biocompatible materials, thepresent invention could be manufactured using this alternate method, asopposed to the layer-based method described above. According to theEFAB™ micromanufacturing process, a patterned layer of sacrificialmaterial is deposited on a substrate, a blanket layer of structuralmaterial is deposited on the patterned layer, and then the resultinglayer is planarized; successive layers are formed in this fashion, andthe sacrificial material is released. To include an embedded structureor device using EFAB™ technology such as optical fibers, some additionalsteps are needed in addition to the standard process. For example, aftera base structure is formed and the sacrificial material is released, anoptical fiber is embedded, the next patterned layer of sacrificialmaterial is deposited on top, a blanket layer of structural material isdeposited on the patterned layer, and then the resulting layer, withfiber optics underneath, is planarized. One skilled in the art ofmicrofabrication will appreciate that although the use of laminatedglass layers is preferred with present technology, the making of theinvention is not limited to a single technology of manufacture. Indeed,any suitable technology for arriving at a relevant 3-dimensionalstructure may be used to fabricate the invention.

Those skilled in their respective arts will appreciate that theinvention represents a highly interdisciplinary technology and isamenable to any number of design considerations.

8. Other Embodiments

Up to this point in the disclosure, embodiments of a side-ventedmicrocradle have been described that translate a horizontal motion offluid into a vertical motion past the patient, so that the patient isventilated vertically. However, referring to FIG. 94A, rather thanproviding side ports for a fluid flow that translates into a verticalventilation of the patient, side ports 219 can be used to ventilate thepatient horizontally instead. Referring to FIGS. 94A-E, FIG. 94A showsan FCA 220 containing a side-vented microcradle 221 that is covered by amicrodrop 222. An optional retaining structure 223, such as a picketfence structure, may be included to retain the patient P in a morelocalized area, while at the same time permitting a flow of fluidthrough the retaining structure 223. Ventilation may occur in the formof flow in a single direction, in alternating directions, to-and-fro, orusing a back-and-forth rocking motion to rock the egg of the patient Pback-and-forth. Note that an intake of fluidic ventilation must equal anoutput of fluidic ventilation in order to maintain a constant volume ofthe droplet 222.

Referring to FIG. 94B, which is a side cross-sectional view taken alonga line 224 with respect to FIG. 94A, the patient P is horizontallyventilated in the microcradle 221 via side ports 219. Note the open top2 and clear bottom 3 of the present invention. Unlike the art of Beebeet al (U.S. Pat. No. 6,193,647), which includes a covered well orcovered channels in which to rock an embryo back and forth using amicrofluidic flow, the open top 2 arrangement of the present inventionenables easy access to the patient P while allowing him or her to remaincontinuously subject to an ambient pressure rather than to a pressureused to urge fluid to flow.

Referring to FIGS. 94C-E, which are side cross-sectional views takenalong a line 225 (flooring layers omitted) with respect to FIG. 94A,side ventilation ports 219 may be formed by channels etched in (226) orthrough (227) layers 228. Referring to FIG. 94E, a top layer 229 capsoff channels 227 etched through layers 228. Note that the patient P willexperience the microfluidic benefits of laminar flow in thisarrangement.

Beebe et al (U.S. Pat. No. 6,695,765) teach a network of coveredchannels in which to house an embryo for purposes including transport toanalysis stations. However, a great liability of their teaching is thatembryos are not available for easy access while traveling inside thecovered channels and, moreover, they are subject to a pressure used tourge fluid to flow rather than an ambient pressure of the environment.

Referring to FIGS. 95A-C, the invention overcomes these liabilities withan embodiment of a side-vented microcradle known as a “micro-canal”.Referring to FIG. 95A, the micro-canal is formed by a microchannel 230with an open top 2 and clear bottom, which houses a patient P in a fluidincubation medium M. As shown in FIGS. 95A-B, a hydrophobic coating 231may be optionally applied to a surface surrounding the micro-canal(microchannel) 230 and a layer of embryo tested mineral oil 232 or otherhydrophobic and biocompatible liquid is placed as a cover over theincubation medium M.

Referring to FIG. 95B, an FCA device 233 or other equipment may beplaced above the patient P in the micro-canal 230 as shown, or below orat the sides. Those skilled in the art will appreciate that such adevice 233 will provide means to perform such exemplary functions asthose of a gate, filter, microsurgical device, optical device,micromanipulator, sensor, transfer catheter docking station, treatmentstation, analysis station, and so on.

Referring to FIG. 95C, which is a particular example of the generalembodiment of an FCA according to the invention shown in FIG. 5, an FCA400 populated with any number of components 27 and micro-canal devices233 cradles a patient P in a micro-canal 230, which serves as a greatlyelongated version of a side-vented microcradle 26. In general, thepatient P will be ventilated with fluid medium M by ventilation ports onthe sides of the micro-canal 230, including by ventilation ports locatedat the ends 234 or along the sides 235 of the micro-canal 230.Ventilation may also be provided via a vented flooring or from above.

Such ports may be used to cause the patient P to transit in the canal bymeans of a fluid flow. Accordingly, the patient P may be moved from onemicro-canal device 233 to another as desired. A circular micro-canal ornetwork of micro-canals may also be employed. Flow in a network ofmicro-canals can be controlled by gates and fluid flow. A recirculatingpump is preferred to maintain an equal inflow and outflow of fluidbetween ventilation ports exhibiting opposing fluid flow. Fluid flow andpatient transit may occur in a micro-canal 230 in a desired manner,including to-and-fro, pulsating, bidirectional, unidirectional, and soon. The course or circuit traveled by the micro-canal 230 may imitate insome respects the course of the human fallopian tube. Spillways andspillway sensors may be employed to help ensure fluid balance ismaintained in the micro-canals, including an optical sensor as shown inFIG. 93 (since overflow will cause a bulging meniscus).

An optical device may follow the patient P on a track (e.g., under themicro-canal) for visualization and thermography as the patient P travelsthe course of the micro-canal 230. Alternatively, an FCA 400 on amotorized x-y stage subject to computer controls may be made to scrollover optical or other equipment in response to patient P movement alongthe course of the micro-canal 230. A computer can be used to performtracking functions digitally.

Oil 232 or other substance, if used to cover the fluid medium M in themicro-canal 230, should not interfere with optics. Although amicro-canal 230 with vertical sides has been shown, sloped or curvedsides are also possible.

Importantly, an FCA containing a micro-canal enjoys the same benefits ofa temperature bath according to the invention as do other side-ventedmicrocradle embodiments.

Referring to FIGS. 96A-B, a minimalist embodiment of a side-ventedmicrocradle 236 with an open top 2 and clear bottom 3 comprises aventilation port 237 and a retaining structure 238 to localize thepatient P. Referring to FIG. 96B, the ventilation port 237 providesventilation from the sides. In FIG. 96B a retaining structure is notshown. However, it is understood that if the retaining structureprovides bidirectional localization for the patient P, then a flowto-and-fro can be provided via the ventilation port 237. In contrast,FIG. 96A shows a retaining structure 238 providing unidirectionallocalization; note that a flooring port (drain) 239 may be employed todrain fluid away.

Referring to FIG. 97, a digital microfluidic system employingself-scooting circuitry according to the invention can be made to urge acontinuous train (or stream) of droplets (or fluid cells) separated andspaced apart by a filler medium. This provides a very powerful hydraulicarrangement capable of pumping fluid with great force. But there is alimitation as to how close together the droplets can be spaced.

To move a droplet by electrowetting a leading control electrode (orcontrol surface) must be turned on and a trailing control electrode mustbe turned off. Accordingly, referring to FIG. 97, for a leading droplet240-1 to move, a control electrode CE(3) at the trailing edge of thedroplet 240-1 must be turned off. Similarly, in order for a trailingdroplet 240-2 to move, a control electrode CE(2) at the leading edge ofthe droplet 240-2 must be turned on. Since an electrode cannot be on andoff at the same time, the leading edge of one droplet 240-2 cannot sharea control electrode in common with the trailing edge of another droplet240-1. Consequently, enough filler medium 241 must be provided toseparate droplets with enough spacing so that they do not share acontrol electrode in common between their respective leading andtrailing edges.

However, in some cases it may be desirable to space conductive fluiddroplets (or fluid cells) even more closely. Referring to FIGS. 98A-B, asomewhat peculiar charged electrowetting arrangement employingself-scooting circuitry is provided to satisfy this goal according tothe invention. Referring to FIG. 98A, a train of fluid cells composed ofa polar, conducting liquid 242 (e.g., aqueous) are separated by animmiscible, non-conducting liquid 243 (e.g., oil) that serves as afiller medium. As shown, adjacent fluid cells 242 are maintained withopposite charge polarity. Referring to FIGS. 98A-B, inner 244 and outer245 layers of control electrodes urge the train of fluid cells 242 inthe manner of charged electrowetting, as shown. Note that adjacentcontrol electrodes in a given layer have opposite polarity with respectto each other and with respect to the fluid cells 242 they are leading.

The length of two fluid cells 242 plus the filler medium 243 in betweenestablishes what may be called the wavelength λ of the arrangement.Charges in the control electrodes layers 244-245 propagate in the samedirection with the same wavelength λ. Quarter-wavelength nodes 246 havebeen inscribed on the drawing of FIGS. 98A-99 for convenience. Thepolarity of a given fluid cell 242 remains constant as it travels. Innerand outer layers of control electrodes 244-245 alternate in being turnedon and off as the fluid cells 242 travel one-quarter wavelength. Eachtime a given control electrode layer 244-245 is turned on and off,individual control electrodes alternate (advance) in polarity, so thatcharges in the control electrodes lead opposite charges in the fluidcells 242. Charges in the fluid cells 242 would come to rest withrespect to charges in either control electrode layer 244-245 at a phasedifference of ½λ; so, to keep the fluid cells 242 flowing constantly thecontrol electrodes propagate their charges with a leading phasedifference of ¾λ as they alternate back and forth between layers244-245.

Referring to FIG. 99, a slightly different arrangement employs only onelayer of control electrodes 247. Trailing control electrodes 248 may beturned off when appropriate; control electrodes may be further dividedas desired so that trailing control electrodes may be turned off more orless continuously.

FIGS. 98A-B and 99 show what are here called lamellae pumps since thefiller medium 243 forms a lamella (thin plate) between fluid cells.

For comparison, consider the approximate size of one droplet (fluidcell) plus the size of a filler medium used between droplets in terms ofa corresponding number of control electrodes needed to cover anassociated amount of space. With respect to FIG. 97, approximately threecontrol electrodes are needed; with respect to FIG. 99, two controlelectrodes are needed; and, with respect to FIGS. 98A-B, only onecontrol electrode is needed. Accordingly, the arrangement of FIGS. 98A-Bis found to be the most conservative in this respect.

Referring to FIGS. 98A-99, ideally the fluid cells 242 are endowed witha net charge that is preserved at least for some time by electricalisolation, and the self-scooting circuitry needed to control the controlelectrodes 244-245, 247 is controlled by an optical sensing of thepassing of fluid cells 242 or filler medium 243. Magnetic orelectrostatic sensing may replace optical sensing. An exemplary means ofmagnetic sensing may include an implementation of the giantmagnetoresistive (GMR) effect. However, in many cases the passing offluid cells 242 or filler medium 243 will have to be electronicallysensed, and charges on fluid cells 242 will need to be constantlymaintained by means of charging electrodes.

Note that although a net charge is placed on a fluid cell 242, thecharge is nevertheless balanced by electrolytic capacitance with respectto the control electrodes, in contrast to a purely static net charge.Still, without good electrical isolation, charge may leak.

Referring to FIG. 100, at one or more nodes 246, in place of an opticalmeans, a pair of switching electrodes 249-250 senses the passing of theconducting fluid medium 242 and the non-conducting filler medium 243 asthe switching electrodes 249-250 form an open and closed liquid contactswitch. At each node 246 a charging electrode 251 placed in contact withthe passing fluid maintains an electrolytic charge on fluid cells 242using nominal self-scooting circuitry, which in turn is controlled by anexemplary means of sensing the passing of the fluid cells 242, such asthe above-stated electronic or optical means. The switching and chargingelectrodes 249-251 must be sized smaller than the width of the fillermedium 243, as shown, so that electrical contact is not made betweenadjacent fluid cells 242.

In a microfluidic system, fluid cells are partitioned by injectingimmiscible media in an alternating series. A computer-readable “barcode” series is formed by spacing the alternating media in long versusshort amounts over a given region in a train of fluid cells. Media ofdifferent optical properties may also be employed (e.g., light vs.dark). The passage of the bar code region over a sensor may be used toprovide information for the control and operation of a microfluidicsystem. For example, electrowetting voltages can be temporarily turnedoff in response to recognition of a given bar code sequence, so thatcertain fluid cells in a continuous train of fluid cells are not exposedto voltages or charges as they pass. Similarly, bar code regions writtenamong leading fluid cells can be used to encode information about thecontents of trailing fluid cells, e.g., for routing purposes.Alternatively, bar codes may be written in fluid cells on a pacer track,or on a separate bar code or reader track.

In general, any method of information science can be used in practice towrite bar codes using a series of fluid cells. For example, fluid cellsof equal length, but of low versus high electrical conductivity, can beused to code for 0 and 1 respectively. Clear versus opaque, high versuslow pH, and other binary contrasts may also be employed.

In general, fluid cells manipulated by digital or analog microfluidicsmay contain various chemical contents. They may also contain cells,sperm, viruses, particles, and so on, which in turn may be subjected tovarious processes, such as transport, sorting, flow cytometry,treatment, handling, and so on.

It may be noted that a micro-canal serves in effect as an elongatedfluid cell for housing a patient, which in turn may be subject toadditional partitioning.

9. Examples of Human Medical Use

The purpose of a prenidial incubator according to the invention is tosustain the life of a premature infant during life before implantation.In other words, the invention enables us to extend our care for preterminfants to life before implantation.

The invention finds use whenever an infant requires incubation outsidethe maternal body during life before implantation.

Examples of medically acceptable uses include a monitoring and care ofthe infant after in vitro fertilization with transfer to the infant'sbiological mother, rescue of the infant after maternal demise withtransfer to an adoptive mother, separation of mother and child withtransfer to a surrogate mother in cases where conception has beendetected yet where continued pregnancy is not possible with thebiological mother, and temporary separation of mother and child withtransfer back to the biological mother in cases where the motherrequires treatments that cannot be safely performed with her infantpresent. It is contemplated that as technology advances it will somedaybe possible to detect a condition of the infant in the maternal bodyduring life before implantation, such that rescue and temporaryincubator involvement will then be indicated for the infant in caseswhere the condition merits it.

The invention enables an infant to sustain meaningful life outside themother's womb from the time of creation until transfer becomes necessaryfor implantation.

10. Other Uses

Because the invention has the clear moral status of an infant incubator,it cannot be used, advertised, or exploited in any manner that iscontrary to patient rights. However, based on the level of technologythat has gone into the making of the invention, it cannot be denied thatthe invention is amenable to a variety of non-human uses as well. Infact, some of these uses may even serve to benefit the care of prenidialinfants in the form of useful research.

Those skilled in the arts of veterinary medicine and microbiology willappreciate that obvious modifications of the invention are amenable tonon-human use. Exemplary mammalian uses include murine, porcine, bovine,and equine use. An exemplary use in the field of developmental marinebiology includes the study of echinoderm development. Those skilled inthe art of electrophysiology will appreciate that modifications of thepresent invention, which include the advantages of microfluidicscombined with an open top arrangement, are superior to the prior art ofperfusion chambers and closed-top microfluidic arrangements.

For example, the invention may be modified to accommodate a Xenopusoocyte.

11. Principles of Prenidial Thermoregulation

Historically, an accurate taking of a patient's temperature proved to bethe gateway to modern medicine. Today, it goes without saying that noclinical practice is more basic to modern medicine than the taking of apatient's temperature. It should also be understood that the keyingredient of an incubator system is found in its ability tothermoregulate the patient. However, modern medicine has been slow tocatch onto the latter understanding.

The nature of the problem of thermoregulating a patient in an incubatoris actually more complicated than may be initially appreciated. Forexample, pediatric historian Thomas E. Cone, Jr. recounts shortcomingsin the history of neonatal incubation by teaching as follows: “Earlychick incubators in this country failed because their users maintained aconstant ambient temperature during the entire 21-day period ofhatching. Eventually they learned the Egyptian secret; namely, that asthe chicks' endogenous [internal, bodily] heat production increased [dueto increased energy metabolism based on growth], the ambient temperaturewithin the incubator needed to be reduced accordingly. The incubation ofhuman infants also stumbled for many years because physicians did notappreciate that environmental temperatures should be reduced asendogenous heat production increased [with growth due to increasedenergy metabolism].” (Cone T. E. Jr. History of the Care and Feeding ofthe Premature Infant. Boston: Little, Brown, 1985. pp. 21-22) Thisimportant and substantially correct teaching is quoted in texts onneonatal incubation today.

Yet the nature of thermoregulation in an incubator system is complexenough that even Cone proves to be befuddled on the subject. For itturns out that modern chick incubators maintain a constant ambienttemperature during the entire period from fertilization to hatching!Since Cone's teaching is correct in context, how can this contradictionbe explained?

Human and animal organisms may be categorized either as thermoregulators(like birds, mammals, and humans) that rely on a given body temperaturefor healthy operation or as thermoconformers (like fish and reptiles)that allow body temperature to change with the temperature of theenvironment. What this implies, for example, is that a chick embryogrowing in an egg actually prefers a certain body temperature.

Neglecting for the moment the additional topic of heat exchange due toradiant energy sources, as well as non-applicable topics such as coolingby evaporation of sweat, the body temperature will be determined by acontrast between bodily heat production, the ambient environmentaltemperature, and one other factor: dissipation. Notably, it is adifference in heat dissipation that explains the contradiction behindCone's teaching.

Egyptian incubators kept poultry eggs in heap-like piles offering poordissipation of heat. Consequently, as internal heat productionincreased, the ambient temperature of the environment needed todecrease. In contrast, a modern chick incubator spaces eggs apart ontrays and the air is allowed to circulate. In this manner, heatdissipation is favorable enough that the ambient temperature does notneed to be lowered to accommodate increases in the chicks' internal heatproduction. However, after hatching modern hatcheries lower the ambienttemperature by about one degree Fahrenheit per day until ordinarytemperatures are reached; otherwise the hatchlings will becomeoverheated due to increasing endogenous heat production, given that thewarmth of their bodies is now insulated by downy feathers, which limitsheat dissipation. Note that incubation is no longer necessary once thechicks are able to produce enough heat to sustain the proper bodytemperature at ordinary temperatures of the environment.

Cone recounts an important observation regarding the Egyptianmethodology, namely, they would place an egg against the sensitive skinof the eyelid in order to determine its temperature. In this way, theyknew from experience whether the egg was too cold or too hot.Accordingly, they would change the temperature of the incubatorenvironment. Importantly, their methodology was correct in that theychanged the ambient temperature of their incubators in response tofeedback from readings of body temperature. In contrast, earlypractitioners of neonatal care made the terrible blunder of monitoringonly the temperature of the incubator, which they endeavored to keepconstant. In this regard, Cone's teaching could be clarified somewhat inthat the major blunder of early practitioners was found morespecifically in a failure to monitor the patient's own temperature, asopposed to the temperature inside the incubator microenvironment. For,it is from a monitoring of the patient's personal body temperature thatthe need to alter the thermal parameters of the environment can beinferred.

Unfortunately, the mistakes of the past have been stubbornly repeated bypractitioners of what has been called in vitro fertilization. Indefiance of my teaching in U.S. Pat. No. 6,694,175, they continue tomonitor only the temperatures of their incubators, and not patienttemperature itself. For example, at Boston IVF (Brookline, Mass.), theonly fertility clinic affiliated with Harvard Medical School, chiefembryologist C. Brent Barrett responded to my teaching as follows: “Theembryos that we incubate are microscopic in size and therefore, there isno difference in the temperature of the interior of the incubator andthe embryo. We constantly monitor the temperature of our incubators andhave conducted numerous studies to ensure that we maintain an optimaltemperature for the embryos.” (Personal communication. Jul. 12, 2004)Note his focus on the temperature inside the incubator combined with hisunskilled assumption that there is no difference between thattemperature and the temperatures of the embryos.

It is known from the science of thermodynamics that heat transfer bythermal contact implies a thermal gradient from higher to lowertemperature. Put another way, heat flows downhill. Therefore, in termsof heat transfer by thermal contact, at steady-state equilibrium eventhe temperature of a thermoconformer must be at a higher temperaturethan the surrounding environment! In other words, because the organismproduces heat, and the heat must be dissipated into the surroundingenvironment, a temperature gradient must exist. Put another way, if thebody and the environment were at the same temperature, the bodytemperature would rise as heat is produced internally, and at that pointa net thermal exchange from higher to lower temperature would occur. Therise in body temperature would then stop once a steady-state equilibriumis achieved in terms of heat dissipation. In this sense, although thetemperature of a thermoconformer changes with the temperature of thesurrounding environment, it does not match it exactly since the bodyproduces heat and heat dissipation by thermal contact implies a thermalgradient from higher to lower temperature. Understandably, this teachingapplies equally to microscopic human embryos, being based on soundthermodynamic principles.

Today, an understanding of these principles is generally a commonplaceexperience. For example, a nurse relies on a patient thermometer, ratherthan on room thermostat readings. A chef relies on a meat thermometer,rather than on thermostat readings of oven temperature. Unlike practicesof old, in which incubator temperature readings were mistakenly reliedupon, today neonatologists distinctly monitor a neonate's bodytemperature without confusion. Even Albert Einstein believed that thelaws thermodynamics always apply, including in the outer space world ofrelativity!

It makes sense to believe that these laws also apply in the world of apetri dish.

Referring to FIG. 101, a prenidial infant P is depicted in reference topoints A, B, and C respectively located inside, on the outer surface,and at a distance of a few microns away from the outer surface of thebody. FIGS. 102A-103B graph temperature as a function of distance fromthe body surface. For the purposes here, the temperature measured on theouter surface of the body is treated as patient temperature, T_(p).

In the graphs of FIGS. 102A-103B, optimal body temperature T_(o) for thepatient P is the same in each case. FIGS. 102A-B illustrate a differencein heat dissipation regarding the rate at which heat produced by thebaby's body P is carried away into the surrounding environment. FIG.102A represents a case of lower heat dissipation than that of FIG. 102B.In both of these figures, however, the temperature of the baby P happensto be optimal; in other words, T_(p)=T_(o). In the case of FIG. 102Aheat is dissipated into the surrounding environment more slowly than itis in the case of FIG. 102B. For example, fluid ventilating the patientP may be flowing at a faster rate in the case of FIG. 102B than it is inthe case of FIG. 102A, thus providing for a greater rate of heatdissipation. All else being equal, in order to establish the optimalpatient temperature condition, T_(p)=T_(o), the ambient temperatureT_(a1) associated with the case of lower heat dissipation (FIG. 102A)will have to be lower than the ambient temperature T_(a2) associatedwith the case of higher heat dissipation (FIG. 102B).

All else being equal, the reason for the difference is that a case ofpoorer heat dissipation (e.g., a stagnant fluid medium) requires a lowerpull-down temperature to allow for the same body temperature atsteady-state equilibrium as does a case of improved heat dissipation. Anappreciation of this difference is very important because it teaches usthat an optimal ambient temperature does not regard an absolute setting;instead, it is a variable quantity that depends on factors such as heatdissipation.

Looked at in another way, suppose the case of FIG. 102A regards astagnant fluid medium and the case of FIG. 102B regards a fluid mediumflowing over the patient P at a rate of X microns per second. Thisimplies that when increasing the rate of flow from 0 to X, the ambienttemperature of the fluid medium must be raised from T_(a1) to T_(a2) inorder to maintain the optimal temperature T_(o) for the patient P.Otherwise the patient P will be chilled, as in a draft, by the fluidflowing at the cooler temperature. In other words, unless the ambienttemperature is raised in response to an increase in flow-related heatdissipation, patient temperature will fall below what is optimal for thepatient P.

Conscious of this critical problem, the parent application (Ser. No.10/908,861) teaches the need to monitor patient temperature in responseto flow and other parameters, so that feedback controls can be obtainedfor the sake of proper thermoregulation.

FIGS. 103A-B show graphs for cases in which patients are overheated(FIG. 103A) or chilled (FIG. 103B). In the case of an overheatedpatient, the patient's temperature is above the optimal bodytemperature; in other words, T_(p)>T_(o). In the case of a chilledpatient, the patient's temperature is below the optimal bodytemperature; in other words, T_(p)<T_(o). FIG. 104 shows a flow chartoutlining a variety of responses that can be taken to bring a patient'stemperature to normal when he or she is overheated or chilled. Referringto FIG. 104, to cool an overheated patient we can lower the ambienttemperature, increase the dissipation of heat (e.g., by increasing therate of fluid flow past the patient), decrease radiant heating, or lowerendogenous heat production. A decrease in radiant heating can include adecrease in heating from radiant heat sources (e.g., heat lamps) as wellas allowing for an increase in radiant heat losses from the patient tothe surroundings. Conversely, to warm a chilled patient we can raise theambient temperature, decrease the dissipation of heat (e.g., bydecreasing the rate of fluid flow past the patient), increase radiantheating, or raise endogenous heat production. An increase in radiantheating can include an increase in heating from radiant heat sources(e.g., heat lamps) as well as preventing radiant heat losses from thepatient to the surroundings.

Consider a unit change in endogenous heat production, meaning, a changein the amount of heat produced by the patient per unit time. All elsebeing equal, if the patient increases her or his heat production, bodytemperature will rise; conversely, if the patient decreases heatproduction, body temperature will fall. However, in going from one stateof steady-state equilibrium to another, referring to FIGS. 102A-B, thenet change in patient temperature ΔT_(p) per unit change in heatproduction will depend on the rate at which heat is dissipated into thesurroundings; more specifically, the change in patient temperatureΔT_(p) observed per unit change in heat production will be lower in acase of greater heat dissipation (FIG. 102B) than it will in a case ofless heat dissipation (FIG. 102A).

In this sense, increasing heat dissipation will indirectly have somewhatof a buffering effect on temperature stability. But unnecessary relianceon this effect threatens to lead to an unsophisticated result. Forexample, it is clear that setting the ambient temperature of the fluidmedium equal to the optimal surface body temperature of the patient andflowing it past the patient at a high enough rate will have the effectof forcing the patient's surface temperature to the optimal temperature,by virtue of heat dissipation, regardless of variations in endogenousheat production. In fact, it might be tempting to think that thisapproach would obviate the need to monitor patient temperature at all.However, such thinking turns out to be non-subtle. First, independenceof the flow rate variable is sacrificed; one problem with this is thatbeneficial substances produced endogenously might be whisked away toosoon at a rate of flow needed to buffer temperature completely; althoughthe rate of flow might be minimized in this respect, an element of lossof control over the variable will still be evident. Second, thisapproach only serves to buffer the patient's surface temperature;referring to FIG. 101, dissipation of heat internally from a point Ainside the body of the patient P to a point on the surface B is also anissue; factors affecting the rate of interior heat dissipation in thisrespect can include insulation 252 surrounding the patient P, such as ashell, corona cells, or spacesuit; this can be a particular problem inthat a rate of heat dissipation provided by fluid flow will notnecessarily be evenly distributed over the patient's surface; forexample, referring to FIGS. 48B-C & E, the patient's formal body may befacing away from the course of fluid flow; thus, flow and, hence, aneffect of heat dissipation can be uneven; the unevenness in turn can bemagnified by the rate of flow itself. Third, forcing a patient'stemperature to a set value will mask important indicators of her or hishealth status.

The problem of masking indicators of patient health status by exercisingtotal control over thermoregulation parameters is a well known problemregarding the care of neonates in incubators in the field of neonatologytoday. For this reason, it is necessary to allow the patient to exhibitdetectable changes in temperature, since these changes can be a sign ofchanging health status. For this reason, generally speaking, some changein temperature ΔT_(p) should be allowed per unit change in patient heatproduction; in this way, the change may be noted and incubationprotocols can be adapted to respond accordingly. In other words, eventhough the response may be, for example, to restore the patient tooptimal temperature, noting the change itself can provide importantinformation that would otherwise be masked.

For example, if the patient's surface temperature is forced to the sametemperature as the fluid bathing the patient using a high rate of flow,then even if the surface body temperature is being monitored it willstill not be possible to distinguish a healthy patient from a deceasedpatient on the basis of body temperature alone. In contrast, if theambient temperature of the fluid medium is less than optimal bodytemperature, which implies a lower rate of heat dissipation, thendeceased patients will exhibit the ambient temperature of the fluidwhereas a healthy patient will exhibit a higher temperature due to heror his heat production.

An incubator for babies should provide a control of body temperaturewithout masking patient health status and without subjecting the patientto inappropriate rates of flow. For example, to produce ventilation onpar with what is experienced naturally in the human fallopian tube, itis evident from natural transit rates that flow rates past the patient'sbody will be extremely subtle. Thus, using high flow rates to maintainpatient temperature will conflict with this subtlety.

According to the invention, basic control of patient temperature isprovided in view of feedback from patient temperature readings based onan adaptable setting of the ambient temperature of the fluid bathing thepatient in contrast to a rate of heat dissipation provided by fluidflow; consequently, there is no need to crudely force the patient to agiven temperature since proper monitoring and control ensures therequisite adaptability. However, although this level of basic controlmay suffice, there are added advantages to employing radiant heat toprovide a fine control of patient temperature in addition to the basiccontrols afforded by the invention. Most important is that radiant heatsources can be quickly adjusted or relaxed, in contrast to changes ofambient temperature or the fluid flow rate. Relaxation techniques, alsoknown as perturbation techniques, enable information about a system tobe gathered as the system proceeds to a new state of equilibrium, or newstate of steady-state equilibrium, after a system variable (orparameter) is relaxed or perturbed.

For example, consider a patient whose body temperature is maintained byinternal heat production, ambient temperature, and radiant heat. Whenthe radiant heat source is turned off, the patient will proceed to alower temperature. The time taken to reach the new temperature can beplotted as a function of temperature, and this curve can yieldinformation about individual characteristics of the patient. A similarapproach can be taken regarding other parameters of incubation.Relaxation techniques form an important part in the sciences, particularphysical chemistry. Thus, it is contemplated that research in this areawill help to elucidate indicators of patient health status as well asindividualized determinations of optimal settings.

Because past practices were incompetent, it should be reflected that anumber of changes will need to be made to incubation protocols inaddition to refining the technology of incubator controls. For example,consider an incubator system in which patients are likely to becomeoverheated due to increases in their own metabolic activity. In thisunfortunate scenario, a formula for the baby's fluid incubation mediumthat suppresses metabolic activity may actually yield higher survivalrates than an optimal formula! For this reason, the present invention isan enabling technology, along with U.S. Pat. No. 6,694,175 and theparent application (Ser. No. 10/908,861), because together theseteachings enable proper control of thermoregulation and ventilation.

In other words, because thermoregulation and ventilation establish theuniversal basis of a competent incubator system, having a competentmeans of thermoregulation and ventilation enables other parameters ofincubation to be determined in a competent light. In contrast, theperformance characteristics of an incompetent means will leave usguessing as to whether given protocols are truly advantageous or merelyadvantageous in an inferior light.

Prenids are to be regarded as being especially delicate compared toother babies insofar as their thermal sensitivities and requirements areconcerned. For example, they appear to be especially susceptible tocytogenetic abnormalities induced by improper thermoregulation, oftenleading to lethal mosaicism. Accordingly, it is all too easy tounderstand why prior art practices have been disastrous and whytremendous medical and scientific reform is needed.

In conclusion, taking care of babies from the moment God has createdthem is a deeply religious form of endeavor. It therefore commands ourhonor and seriousness. The advent of prenidial care not only representsa great milestone for medicine, but, like neonatal care before it, alsoan immeasurable gift for the human family. Accordingly, those skilled inthe art of health care will appreciate that an incubator for babiesduring prenidial development demands subtlety, sophistication, andtechnological elegance, as taught by the present invention.

What is claimed is:
 1. In an incubator for babies before implantation,wherein a means of patient thermometry detects an actual patienttemperature and which said temperature can differ from an ambienttemperature of incubation liquid media ventilating the patient, a methodof thermoregulation for a human embryo or hatchling within theincubator, comprising steps of: (a) detecting the actual patienttemperature and comparing it to a predetermined value for an optimalpatient temperature; (b) when the actual patient temperature is lowerthan the optimal patient temperature, warming the patient by means ofthermal controls selected from the group consisting of raising theambient temperature, decreasing a dissipation of endogenous heat,increasing a radiant heating of the patient, and raising endogenous heatproduction; and, (c) when the actual patient temperature is higher thanthe optimal patient temperature, cooling the patient by means of thermalcontrols selected from the group consisting of lowering the ambienttemperature, increasing a dissipation of endogenous heat, decreasing aradiant heating of the patient, and lowering endogenous heat production.2. In an incubator for babies before implantation, wherein amicrofluidic means controls a rate of flow of incubation liquid mediapassing over in fluidic contact with a human embryo or hatchling, amethod of patient thermoregulation within the incubator, comprisingsteps of: (a) maintaining an ambient temperature of the incubationliquid media in fluidic contact with the patient, wherein the ambienttemperature is preset to a temperature lower than an optimal patienttemperature; (b) detecting an actual patient temperature by means ofpatient thermometry and which said actual patient temperature can differfrom the ambient temperature; (c) comparing the actual patienttemperature to the optimal patient temperature; (d) when the actualpatient temperature is higher than the optimal patient temperature,signaling the microfluidic means to increase the rate of flow, wherebyconsequent increases in flow-related dissipation of endogenous patientheat lower the actual patient temperature closer to the optimal patienttemperature; (e) when the actual patient temperature is lower than theoptimal patient temperature, signaling the microfluidic means todecrease the rate of flow, whereby consequent decreases in flow-relateddissipation of endogenous patient heat raise the actual patienttemperature closer to the optimal patient temperature; (f) repeatingsteps (b) through (e) until the actual patient temperature is equal tothe optimal patient temperature or until the rate of flow is higher orlower than a predetermined range of acceptable flow rates; (g) when step(d) results in the rate of flow which is higher than the predeterminedacceptable range, lowering the ambient temperature; and, (h) when step(e) results in the rate of flow which is lower than the predeterminedacceptable range or zero, raising the ambient temperature.
 3. The methodof claim 2, wherein the means of patient thermometry comprises infraredmicrothermography.
 4. The method of claim 3, wherein the optimal patienttemperature is 37 degrees Celsius.
 5. The method of claim 1, furtherincluding: (d) when the actual patient temperature is in a state ofequilibrium or steady-state equilibrium, relaxing or perturbing at leastone of the thermal controls and charting a progress of the patient to anew patient temperature.
 6. The method of claim 5, wherein the relaxingor perturbing commences when the actual patient temperature is equal tothe optimal patient temperature.
 7. The method of claim 6, furtherincluding: (e) repeating steps (a) through (c) to restore the patient tothe optimal patient temperature; whereby the patient is maintained atthe optimal patient temperature along with periodic charting of thepatient temperature in response to relaxing or perturbing the thermalcontrols.